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Archive for the 'Uncategorized' Category
Puja & Pragnesh (Part II) – WHC Temple, Washington, DC
As the year comes to a close, many of our summer brides are sharing images from their big day! To start, let's take a glimpse at a family wedding from July. The traditional bright red was paired with an orange, garden green, and fuchsia purple to create a festive ambiance. Wedding day stationery included event signage and programs in the afternoon, and coordinated menu cards, table numbers, escort cards for the reception – all in the coordinated Layla Rue design! We love these detail shots taken by the couples' photographer Sunny Mathur of Photographick Studios.
Ariana & Amardeep – Sanctuary Resort on Camelback Mountain, Paradise Valley, Arizona
What's not to love about Ariana and Amardeep – the fun, creative couple from California will be getting married this fall in Paradise Valley, Arizona. Each of their events have a distinctive theme (and illustrations on the event inserts, to match) – including a cowboy-themed Rehearsal Dinner! We love these photographs of the gorgeous couple, as captured by the very talented Sameer Soorma. Their events are being planned by Arizona based wedding planner Suzanne Taussig.
Teejana & Snehit – Save The Date, Destination Mauritius
Update: We love this airplane charm that Teejana selected to adorn her save the dates!
This has to be one of the most unique save the dates we've done to date. Teejana and Snehit's save the dates featured a world map of cities that their guests will be flying in from to their week long destination wedding events in the beautiful island of Mauritius. New York City, London, Paris, Shanghai, Sydney, Mumbai, and Durban were just a handful of the major cities. When we met with the couple in New York City, they described it as being nothing short of a U.N. gathering. To give this destination save the date an upscale look, the silver boarding pass sleeve was printed in silver foil. We absolutely love their monogram crest – which will be incorporated in every part of their invitation and wedding day stationery.
Varuna & Arnob – Grand Cayman Marriott Beach Resort, Grand Cayman Islands
It was truly a pleasure working with New York City based couple Varuna and Bobby, who are hosting a destination wedding in the Cayman Islands this October! After sifting through samples, they fell in love with the layout and style of Vijay and Kalyani's invitation – which we then customized for them using a deep sapphire blue paper, featuring a large vertical monogram. Our favorite details are the small illustrations on their RSVP card to indicate dietary preferences. Photograph of the happy couple by the Brooklyn Bridge courtesy of their photographer Michael Webber of Studio Webber. Their events are being planned by New York City based wedding planner Sonal J Shah Events.
Aarthi & Stephen – The Fairmont, Pittsburgh, Pennsylvania
Aarthi and Steve will be tying the knot this October at the immaculate Fairmont Hotel in Pittsburgh! Aarthi, who was referred to us by her sister and our past client Preethi, chose a simple square bookfold folio suite in a charcoal gray paired with pearl white inserts and envelopes. The invitation was laid out in clean, simple typography accented with a new paisley design. Photographs coming soon!
Amy & Jatinder – Driftwood Farm, Orient, New York
We love when couples take a base design from our collection and do something completely new with it. When we met Amy and Jimmy earlier this year, Amy fell in love with the design and larger square size of Rachna and Samir's invitation – but wanted the papers and color palette to reflect the nature focused event being hosted at a farm. The resulting invitation was created in her softer color palette of coral, peach, and pink and finished on matted textured ivory and white papers. We absolutely adore the engagement shoot photographs that Amy shared with us, courtesy of Raina Kitt, Indigo Foto!
Modern Indian Wedding – The Rockleigh Country Club, Rockleigh, New Jersey
Update: This invitation was featured in South Asian Bride Magazine's blog, as part of a post on ways to dress up your invitation. Read more here!
Enjoy this sneak peak of S&A's wedding invitation suite. True to a modern Indian wedding, the invitation included several event inserts for the various celebrations and were all finished in gold foil printing on a shimmer ivory-gold textured paper. We absolutely love this suite, and are looking forward to creating the coordinated wedding day stationery! Photograph of the happy couple by their wedding photographer, Kate Connolly of Off BEET Productions.
Exciting News!
We have some very exciting news to share! Last week our collection with stationery e-commerce site Wedding Paper Divas went live, and you may now directly purchase some of our South Asian inspired prints online. Wedding Paper Divas (WPD) is a part of the Tiny Prints and Shutterfly Inc. family of brands. What we love about WPD (and makes them a great partner) is their high attention to detail and superior customer service. If you are currently not in need of wedding stationery, you may even purchase our prints for your personal/thank you stationery, or purchase Eid cards for the upcoming holiday. Later this year you will also be able to purchase Diwali Cards, Christmas/Holiday Party invitations, and New Years Eve celebration invitations designed by ZOYA Couture directly on the TinyPrints site!
Placing an order on WPD is simple, easy, and economical. Simply select your components and start customizing your text. Each invitation suite is accompanied by a back design and can be coordinated with matching RSVP cards, additional event inserts, direction/accommodation cards, save the dates, and thank you notes. All pieces are digitally printed on cardstock papers. Ordering through WPD is a perfect solution for clients looking for simple, elegant invitations. Now through 9/30/2012, you can enjoy 25% off our collection by entering the code "ZOYAVIP" at check out. For those looking for a more luxurious finish customized with textured Italian papers and processed with specialized printing such as foil stamping or blind embossing, our custom design team is also available to work with you!
Krupa & Derik – Save The Date, Scottsdale, Arizona
Derik and Krupa will be getting married this November in Scottsdale, Arizona. Sister of the groom and top wedding planner Sonal Shah requested their save the dates to give guests a flavor of the desert and cactus enamored state. We completed these suites on an earthy green paper and imprinted the save the date information with a mountain landscape image in copper foil. The coordinated envelope was printed with the return address – and this particular example showcases a matching green ink in the calligraphy. The calligraphy on the sample is courtesy of Caren Milman, a calligrapher we highly recommend who is based in Rockville, Maryland.
Anne & Santiago – St. John Baptist Catholic Church, Silver Spring, Maryland
Pin-tastic News – We are now on Pinterest!
We had been pinned, and we didn't even know it. As we were recently browsing a few inspiration boards, lo and behold, we found a few of our own invitations! It was a truly lovely surprise. A few hours later of scanning and pinning all kinds of inspiration – we realized why this is such a useful tool in planning events such as weddings. We are working on updating all blog and site images to be easily pin-friendly, and in the mean time invite you to visit our boards, with a preview of two below!
Vanessa & John – Terranea Resort, Rancho Palos Verdes, California
We recently had the opportunity of working with Vanessa and John, from sunny California, on their wedding invitation suite and menu cards. The happy couple is tying the knot at the Terranea Resort, an ocean front venue in close proximity to Los Angeles. The bride-to-be fell in love with Vijay and Kalyani's double matted invitation, and we worked together to re-create the look in a new color palette and size to coordinate with their events. Ultimately, we decided on a textured ivory gold paper as the base for their suite, with the event information in a midnight blue. To match the golden matting of the invitation, we also created their RSVP envelopes in the same paper, imprinted again in the midnight blue ink. We absolutely love these colors, perfect for a luxury resort weekend of events!
Update: We love this engagement shot of Vanessa and John, shared with us by the happy couple! Photograph by Callaway Gable, a leading celebrity and destination event photographer based in Los Angeles, California.
Puja & Pragnesh – Washington Hebrew Congregation, Washington, DC
We are so excited about an upcoming wedding in the family this July! It also meant complete creative freedom when it came to the stationery. Puja and Pragnesh will be hosting their wedding events at the Wyndham Garden Hotel, Greater Baltimore Hindu Temple, and Washington Hebrew Congregation Center. Puja's only requirement was that the invitations be colorful, festive, and elegant. To accomplish this, we laid out her invitation in the brand new Layla Rue design from our ready to order collection, finished entirely in silver foil printing. Our favorite piece of the suite is her RSVP envelope, which accents the design on the crease of the flap quite perfectly!
Sangeeta & Dino – Shangri-La, Ontario, Canada and Dreams Palm Beach Resort, Dominican Republic
Post coming soon!
Jessica & Rick (Part III) – Corcoran Gallery of Art, Washington, DC
It's here – Jessica and Rick's wedding suite finale! With the wedding just one week away, we are excited to share a glimpse of the final coordinated materials – which include tall menu cards that compliment the wedding invitations, folded wedding programs printed on a textured, shimmer gold paper, escort and place cards (being detailed by their talented calligrapher), cocktail signage, and guestbook notes. Our favorite piece among all was also the most challenging – creating the perfect signature cocktail cards. For Jessica's Sri Lankan inspired drink, The Pearl, we included a watermark of the country's outline. For Rick's cocktail inspired by Utica, we used an old family photograph of the city as the backdrop. Have a look! We can't wait to share professional photographs of the materials against their carefully selected decor at the Corcoran.
Karunya & Srikant – The Waldorf Astoria Hotel, New York, New York
It was truly a pleasure to work with Krishna (sister of the bride), on the wedding day stationery for Karunya and Srikant! The wedding events took place at the breath-taking-ly beautiful Waldorf Astoria Hotel and Intercontinental Hotel in New York City, as planned by their event coordinator Sonal Shah. We hope to soon share photographs of their wedding day stationery against their swanky décor, as captured by their photographer Matei Horvath!
Deepa & Anuj – Cipriani Wall Street, New York, New York
When we met Deepa and Anuj for their initial consultation in New York City, we poured over all different design options – and finally decided on a simplified circular motif that incorporated flourishes and paisleys. One element they two absolutely loved was the bronze seta (lined) shimmer paper with blind embossing, and this is easily our favorite part of the suite too! As no ink is used in this method, it is the perfect way to stand-out in a subtle way.
Girish & Kirti – The Doubletree Hotel, Tarrytown, New York
During our first consultation with Girish, he told us how much he loved The Melia design but that he wanted a unique layout of the materials – something that hasn't been done before. A graphic designer himself, Girish took great care in selecting typography and color palette – and inspired us to create a new single panel paper folio to carry his invitation components. With a unique diagonal cut on each side, the folio has an entirely new look, custom created for the happy couple! Girish and Kirti will host their wedding events at The Double Tree in Tarrytown and Greentree Country Club in New Rochelle, as planned by their event coordinator Sonal Shah.
Elina & Pascal – Akerby Herrgard, Nora, Sweden
Today we are thrilled to be bring you yet another international wedding suite! Earlier this year we worked with Pascal and Elina, who reside in The Netherlands, on their stationery for wedding events being held at the beautiful Akerby Herrgard in Nora, Sweden. Despite the logistical challenges (including a 6 hour time difference and not being able to review papers, fabrics, and print styles in person) the project was a lot of fun. The invitation was laid in a champagne colored silk folio, with a clinched ivory ribbon and embellishment on each door. Inside, all components were screen printed in a midnight blue ink on a frosted gold paper. We learned a great deal from working on this suite – including cultural nuances such as using the verbiage "Tenue de Ville" in place of "Black Tie Optional". The folded menu cards listed each course paired with a different wine in both Swedish and Dutch. Finally, the place cards with the couple's monogram were embellished with Swarovski crystals. We loved working on this invitation suite and wish the happy couple the very best with their wedding this July!


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Kinross Declares Dividend TORONTO, ONTARIO -- (Marketwire) -- 02/13/13 -- Kinross Gold Corporation (TSX:K)(NYSE:KGC) announced today that the Board of Directors has declared a dividend of US$0.08 per common share, payable on March 28, 2013 to shareholders of record at the close of business on March 21, 2013. This dividend qualifies as an "eligible dividend" for Canadian income tax purposes. About Kinross Gold Corporation Kinross is a Canadian-based gold mining company with mines and projects in Brazil, Canada, Chile, Ecuador, Ghana, Mauritania, Russia and the United States, employing approximately 8,000 people worldwide. Kinross maintains listings on the Toronto Stock Exchange (TSX:K) and the New York Stock Exchange (NYSE:KGC). Contacts: Media Contact Steve Mitchell Vice-President, Corporate Communications 416-365-2726 email@example.com Investor Relations Contact Tom Elliott Vice-President, Investor Relations 416-365-3390 email@example.com
Kinross Declares Dividend
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subset: commoncrawl


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Report: Former Sheboygan mayor told police he'd prefer arrest by 'hot' cop
4:45 PM, Mar. 8, 2013
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Report: Former Sheboygan mayor told police he'd prefer arrest by 'hot' cop
The night former Sheboygan mayor Bob Ryan was arrested for drunk driving in Kiel, he told police he?d prefer to be arrested by a ?hot? Manitowoc police officer and mentioned calling in favors from
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Chapel Services
Coming Soon
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Real Talk a Tuesday Night Bible Study
Join other IWC students as we explore the Bible and answer your questions.
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Learn how to "run the race" of faith and grow in your relationship with Jesus Christ. FCA is open to both athletes and non-athletes.
The Pastor to Students is happy to talk with you about your spiritual questions and personal struggles. Drop by the Campus Ministry office (Holland Student Center, 2d floor) or call (319) 385-6398 for an appointment.
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Information about faith-based scholarships is available from the Campus Ministry office.
For information on these or other Campus Ministry events, contact Pastor Ron Bupp, (319) 385-6398 or email email@example.com.


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The Gadsden Panthers boys basketball team beat Oñate 39-17 on Friday at Oñate High School.
Tommy Cano led the Panthers with 11 points while Ralph Martinez scored nine for GHS.
The Panthers bounced back from a recent road loss at Alamogordo to move to 4-1 in District 3-5A play. The Knights are now 0-4 in league.


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We have been considering getting a second horse that both my total beginner husband and 9 yr old step-daughter can ride. I found this horse and am wondering if the price is right. We are going to trial him for a week.
Wondering what you would expect to pay for a horse fitting this description in your area...please remember to give your general location!
14 yr old, AQHA registered gelding, Chilled Champaigne (would love to hear what you think about his lines...I don't know anything about it!)
Palomino, 14.3 hh
Trail ridden for 5 years by total beginner husband, goes through everything.
Used as lesson horse for past year, western and english
Neck reins, responds to leg pressure and bit pressure.
WTC, small jumps
Did well at recent dressage show
No illness/injury in past 5-6 yrs (since she has had him)
Very respectful of rider
Perfect ground manners
Assuming he has decent confo, hard to tell from photos
Watched a video of a 4 yr old riding him...he was very gentle!
And handsome too!


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Chandler MVD - Map, Hours, Contact
Chandler MVD Address
Chandler MVD Hours
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Reviews of Chandler MVDAdd Your Review
her name is carol, the overweight rude woman in desk 4. she was already not in her mood when we sat down in front of her, she talks to us in a rude manner. after my son's paper was put to desk1, the woman in desk1 told me to check the details w/c I did, and I found some error and show it to her. the woman went to carol's desk and came back and told me to wait, when all of a sudden, CAROL, IN A VERY LOUD VOICE, told me, in front of these people waiting, that we're supposed to tell her our zipcode changed. I said we put the correct zipcode in the application paper. then she tells me about our address blah blah... I was so upset, I was so embarrassed, my body started shaking...MY GOSH, SHE BLAMES US FOR THE ERROR SHE MADE....in the first place, it is her job to check on the paper, she was already in a hurry when we sat down in front of her, that's why she made an error. I hope dmv will not hire rude lazy people.


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Tag "odroid"
ODROID On Its Way to Early-Bird Devs
The ODROID Developer Edition package is now available worldwide and has begun deployment to the 300 developers that pre-ordered back in September. Will the ODROID gain any traction in the gaming community and break into the market that the iPod Touch is dominating?
ODROID Pre-Orders Announced for Korean Developers, Limited to 300
A platform development version of the ODROID device was announced for preorder in Korea today. Limited to Korean developers, there only 300 units up for grabs. The ODROID development device
ODROID Demo Gives Best Look at Device Yet [VIDEO]
The fine folks at HardKernel were kind enough to pass along another video showing their ODROID device running an Android app up close. The demo below shows the app RockOn,
ODROID in the Wild, Demo's HDMI Output [VIDEO]
Remember that Android-based entertainment device from a couple of weeks ago called the ODROID? It's back today in the form of a demo video. The clip below shows how Android
Meet the ODROID Entertainment Device
Korean-based Hard Kernel were kind enough to pass along some information about their ODROID Entertainment Device. Packed with great specs, it's definitely a unique looking device. It's pretty obvious from


text: Hi, 1. As we know that pressure is force per unit area but why it is said that pressure is a scalar quantity because force is used and force is a vector quantity. 2. Whether stress is also a scalar quantity? 3. What is basic difference between pressure and stress? Regards, Muhammad Rizwan Khalil


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Originally Posted by ic-racer
I used "daguerreotype" and "tintype" to search for those images. So, I'd place most of very roughly in the 1800s to very early 1900s.
Thanks Dale.
"If you want to be famous, you must do something more badly than anybody in the entire world." Miroslav Tichý


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The meter/timer is quite sophisticated so it must have been made by another company and badged for Polysales, who were a sort of UK version of Spiratone with a smaller range. There catalogue was very interesting.
It's not Rayco style but it might have been made by Melico.
Ironically the outdated Bulgin plugs & sockets & old 3 pin 5 & 15 amp UK mains plugs were far safer and less prone to problems that the current ones, andas for the European ones they are even worse. I've never seen the old UK 3 pin plugs burn out, but it's common with the current ones, and the frlimsy 2/3 pin European ones.
I'd rather have a solid Bulgin plug than the flimsy DIN plugs


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These Playful Science Projects Were Created In Just 24 Hours
The idea of Science Hack Days is simple: gather together a bunch of people to get excited and make things with science in an extremely limited period of time. Here are a few of the projects hacked together in San Francisco in just 24 hours. »


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BLACK FRIDAY ends tonight! Save $25 off $200* Use code: TLBF15
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The memory that's crippling Armenia
100 years after their ancestors were massacred, Armenians want both growth and atonement. Right now, it's hard for them to have both.
Obama to shun saying 'genocide' on 100th anniversary of Armenian massacres
Obama will send Treasury Secretary Jack Lew to Armenia this week.
#Armenia: Kim Kardashian meets foreign leader, tweets selfie
The Armenian government just got a taste of Kardashian diplomacy.
Armenia presidential hopeful shot
A doctor says the candidate was injured by an unidentified gunman in the Armenian capital.
What's at stake as Europeans vote
Here's what's at stake Sunday, as six European countries hold elections.
Obama avoids 'genocide' in statement
The president marks Armenian Remembrance Day, while taking care not to offend Turkey.
Flier fuels fierce debate in Ohio
The controversy surrounding the congressional Armenian genocide resolution has a new battleground.


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LAHORE: The first session of the Punjab Assembly has been convened for June 1 and Governor Punjab Makhdoom Ahmed Mehmood has signed the summary in this regard, Geo News reported.
According to details, newly elected members of the provincial legislature would swear-in in the first session.
Provincial Secretary for Law Justice and Parliamentary Affairs Dr. Abul Hasan Najmi told Geo News that the session of Punjab Assembly has been summoned on June 1 at 10:00 am.
According to sources, Speaker Punjab Assembly Rana Mohammad Iqbal would chair the assembly session, after which separate sessions would be called to elect speaker and deputy speaker of the house.
Punjab Caretaker Chief Minister Najam Sethi had earlier said that CM-in-waiting Shahbaz Sharif would take oath on June 6.


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Mini Guide to Combining Lighting Controls and Emergency Lighting
In many lighting installations the lighting serves a dual purpose, with luminaires operating as normal building lighting in normal conditions, plus in an emergency some of them illuminate to provide escape lighting.
When the luminaires are operating as normal building lighting they will be controlled by a lighting control system.
An effective lighting control system is important for an energy-efficient building and for building regulations approval.
Under emergency conditions the escape lighting must illuminate quickly, reliably and unconditionally. Regular testing of the emergency lighting is also a legal requirement.
This guide outlines designs for a lighting installation that will serve both purposes without compromise. It is divided into two sections, describing:
Self-contained emergency luminaires
– luminaires with an inbuilt emergency power source plus all other components required to provide light in an emergency;
Centrally supplied emergency luminaires
– here the power for multiple emergency luminaires comes from a central emergency power system.
In each section the guide describes normal operation, emergency operation and emergency testing.
This guide concentrates on the use of DALI control gear. This is because:
- energy efficient buildings call for dimming and DALI is the most widely used standard for dimming control;
- the DALI standard is unique in that it includes specific provisions to realise dual-purpose lighting in a vendor-independent manner.
Download your copy opposite
Lighting Controls Mini Guide
This guide describes the main benefits of a lighting control system. It provides an overview only and does not drill down into the technologies or design methodologies used in realising a control system.
This mini guide sets out what can be achieved by deploying a lighting control system.
Don't forget to read the Lighting Controls Guide by Chapter, structured to give you a logical progression through the subject of 'lighting controls'.
If you would like to find out more detail on the design, specification, procurement and installation of a control system, as well as the practical issues and technologies available.
Contact Jo: firstname.lastname@example.com or complete the form opposite.
October 2014
Version 2012


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All Comments
what a gorgeous dick. would love to play with it
Nice demonstration and a beautiful dick.
MMMM i just love these videos, I can't get enough of seeing hot cocks in their different states
Good job. Wish I still had that mojo.
love the soft to hard and different positions. Nice dick
Great looking circumcised cock, soft and hard.
I enjoyed this video. Its nice to see what I call normal
sized guy...Most of these videos the guys soft is bigger than most
hard guys...so its kinda not real. But, you are awesome, shows
real life people who are not 5 inches soft...Right on!!
Damn that is one lucky chair mat. Would love to see more like this you have an awesome body and love the furry arms and legs. I had to check out all your vids since I saw that camera cumshot and had to see more. Thanks so much for sharing.


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Link Time!!!
by Celebrity
Like us on Facebook
- Top 10 most ridiculous things about the Beyonce experience — fourfour
- More of Christina's Marie Claire photoshoot — Egotastic
- Watch Posh strut her stuff on stage, so fierce! — D-Listed
- Welcome new PageSix.com!
- Masi and Hayden's fun FHM outtakes — Pink is the New Blog
- Watch Jessica's Disney World Parade performance — Just Jared
- Ridley Scott's making a Gucci movie — BuzzSugar
- Adriana Lima pictures perfect — cityrag
- Amy Winehouse is crying and probably a little cold — Hollywood Rag
- Is Paris using botox too? — Hollywood Tuna
- Ashley Tisdale gets a nose job — People
- Another reality show might be fake, shocking — IDLYITW
- Unappealing maternity clothes walk-off — MollyGood
- Check out Mary J. Blige's new video — popbytes
Read More Link TimeChristina Aguilera
POPSUGAR, the #1 independent media and technology company for women. Where more than 75 million women go for original, inspirational content that feeds their passions and interests.
aprild 7 years
I thought the Beyonce video was dumb b/c it was dissing her. Her concert was great (I went to one in the summer) and she is one amazing performer! Its a shame when people hate on the talented....... She's great at what she does! She sings well, dances well, and puts on one great show!
jaan_black 7 years
deviated septum....aka coke problem....I almost deviated my septum with that drug which was the main reason I stopped...besides um, I love myself and don't want to die prematurely
epicurean_Essy 7 years
"The Beyonce experience" is soo overwhelming. She looks like she is trying to bring someone from the dead!
Martini-Rossi 7 years
Top 10 most ridiculous things about the Beyonce experience is soo funny! She's such a diva. lmao!
Martini-Rossi 7 years
Top 10 most ridiculous things about the Beyonce experience is soo funny!She's such a diva. lmao!
lovekailua 7 years
ugh that is the best news all day. i despise "a shot at love" just from the super annoying commercials, i'll admit i even watch the hills every week and i HATTTTTTE even the commercials for that tequila girl show, bye bye good riddance!
wicca_08 7 years
I'm afraid Hayden's going to be another 'Lindsay' or 'Britney'
wicca_08 7 years
Victoria looks like a robot.
wicca_08 7 years
Victoria looks like a robot.
wicca_08 7 years
I agree, it seems that Beyonce is performing a ritual dance, haha!
pink_magnetism 7 years
:ROTFL: at the Beyonce video!!LOL at Paris' new lips. I really don't get celebrities who think that's a good look.I wonder what Ashley Tisdale's new nose looks like?
pink_magnetism 7 years
:ROTFL: at the Beyonce video!! LOL at Paris' new lips. I really don't get celebrities who think that's a good look. I wonder what Ashley Tisdale's new nose looks like?
Karma-Co 7 years
Ha ha! Beyonce's crazy! But i love it! Ha ha
leeluvfashion 7 years
haha, beyonce is so into herself! that video was priceless.
cocomademoiselle 7 years
Thanks for posting the "Beyonce Experience" video!!! Very funny!!!
NiceDriveway 7 years
I think the most ridiculous thing about the Beyonce 'experince' is ALL.THE.WIND. ALL.THE.TIME.
NiceDriveway 7 years
I think the most ridiculous thing about the Beyonce 'experince' is ALL.THE.WIND.ALL.THE.TIME.
lily3484 7 years
Christina was so quiet about the pregnancy, now she is baring all. I think all of this is little too much. She asks for all this "privacy" and then runs off taking tasteless naked pregnancy pictures.
jmast 7 years
I think Amy Winehouse looks so much better without makeup...but I wish she would have thrown on a sweatshirt. Girl is a wreck.
KristinDaBomb 7 years
Christina = <3
KristinDaBomb 7 years
Christina = <3
angelina8023 7 years
LMAO at the Beyonce video!! Christina looked better in these shots compared to the orange shots:)
loveyoulots 7 years
I love beyonce she's great and that clip was funny as ever especially the check up on it part but i love the tears, man i loved the whole thing.
musiclvr224 7 years
I am not sure I believe the reason on why Ashley Tisdale got her nose job.
alllacedup 7 years
Oh that Beyonce clip made my stressful day! That is the most ridiculous performance I've ever seen!


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While the genetic transformation of the major cereal crops has become relatively routine, to date only a few reports were published on transgenic triticale, and robust data on T-DNA integration and segregation have not been available in this species.
Here, we present a comprehensive analysis of stable transgenic winter triticale cv. Bogo carrying the selectable marker gene HYGROMYCIN PHOSPHOTRANSFERASE (HPT) and a synthetic green fluorescent protein gene (gfp). Progeny of four independent transgenic plants were comprehensively investigated with regard to the number of integrated T-DNA copies, the number of plant genomic integration loci, the integrity and functionality of individual T-DNA copies, as well as the segregation of transgenes in T1 and T2 generations, which also enabled us to identify homozygous transgenic lines. The truncation of some integrated T-DNAs at their left end along with the occurrence of independent segregation of multiple T-DNAs unintendedly resulted in a single-copy segregant that is selectable marker-free and homozygous for the gfp gene. The heritable expression of gfp driven by the maize UBI-1 promoter was demonstrated by confocal laser scanning microscopy.
The used transformation method is a valuable tool for the genetic engineering of triticale. Here we show that comprehensive molecular analyses are required for the correct interpretation of phenotypic data collected from the transgenic plants.
Keywords:Agrobacterium; Winter triticale (x Triticosecale Wittmack); Sexual transmission; Transgene expression; gfp; Copy number
Triticale, the artificial wheat x rye amphiploid, was created in an attempt to combine the grain quality and productivity of wheat with the superior performance of rye in marginal environments. Since the development of the early hybrids, substantial breeding progress has been made by conventional means. Nevertheless, the global cropping area devoted to triticale remains low, and the bulk of its production is concentrated in central and eastern Europe [1]. Given the prevailing climatic conditions in this region, >90% of the crop is represented by winter (vernalization-requiring) cultivars. Its major end-uses are as feed or fodder, but improvements in its grain milling and bread making quality could allow an extension to its use in human consumption or as an industrial feedstock. A particularly attractive prospect lies in its use as a source of bioenergy, but generating the necessary significant changes to the plant to allow this will probably require genetic modification. Hence there is a need to develop robust protocols for its genetic transformation.
The genetic engineering of triticale is still in its infancy. The direct delivery of DNA into protoplasts has achieved transient transgene expression, but not stable integration [2,3]. Difficulties in regenerating plants from isolated protoplasts have prompted the exploration of more readily regenerable explant materials, among which the immature embryo has proven to be the most promising [4-7]. The earliest reported stable transgenic triticale plants relied on the biolistic treatment of immature embryo scutella [8]. A similar approach targeting haploid embryo-like structures resulted in only the transient expression of the transgene and no adult plants were regenerated [9]. A first applicative approach was published by Doshi et al. (2007) who used the embryo-specific LTP1 promoter to show the effect of the C1 and Bperu maize genes on anthocyanin biosynthesis in triticale [10].
Agrobacterium–mediated gene transfer has proven to be an effective means of transforming each of the major cereal species, including triticale [11-13]. For the latter however, robust data on T-DNA integration and segregation were not provided thus far. Using plants produced by means of a previously published protocol [13], we here provide comprehensive information on transgenic triticale in terms of T-DNA integration, copy number, integrity and inheritance. Moreover, we provide evidence of consistent transgene expression across generations.
Primary transgenic plants
Primary transgenic triticale plants that we had reported on in a previous article presenting protocols for several small grain cereals [13] were subjected in the present study to a comprehensive analysis. In this context, we also provide some complementary information on the elaboration of the transformation protocol previously published. As shown in Table 1, a two or five day period of pre-culturing (prior to exposure to A. tumefaciens) was required to obtain transgenic plants. Moreover, inoculation with Agrobacterium proved successful provided the immature embryos were stacked on filter paper moistened with liquid co-cultivation medium, rather than being submerged in the medium. Table 1 further shows, that transgenic plants were exclusively obtained when barley co-culture medium (BCCM) was used, whereas wheat co-culture medium (WCCM) failed to give rise to transgenics under the same conditions. Likewise, a hypervirulent derivative of A. tumefaciens strain LBA4404 resulted in the delivery of transgenic regenerants, whereas AGL-1 surprisingly did not do so under the conditions tested. The expression of gfp was monitored throughout the transformation process (Figure 1A-J). After three days of co-cultivation with A. tumefaciens, fluorescing foci were visible at the margin of scutella of the immature embryos. After a further 3–6 weeks, GFP accumulation was particularly strong in the rapidly growing portion of the callus (Figure 1F-H), whereas emerging leaves and roots showed fluorescence of lower intensity. The transgenic regenerants confirmed by a PCR-based amplification of a gfp fragment did not show any obvious morphological effect associated with transgenicity.
Table 1. Effect of pre-cultivation, A. tumefaciens strain and co-cultivation medium on the generation of stable transgenic triticale
Figure 1. GFP accumulation in transgenic lines as shown by correlative microscopy using white light (A-E) and excitation by far blue light (F-J). Immature embryos (A, F) three days, (B, G) three weeks and (C, H) six weeks after co-culture with Agrobacterium; (D, I) immature pollen formed by the T0 plant TG5E02 isolated from anthers, collected in 0.4 M mannitol and examined under a fluorescence microscope; (E, J) immature T1 embryos from TG5E03 were germinated on K4N medium and their roots examined 5 days after embryo isolation. Non-transgenic tissue domains and segregants are indicated by arrows in B, C, D and E and their fluorescence microscopic counterparts G, H, I and J, where no or only background fluorescence is seen in these positions. Note that the ratios of transgenic and non-transgenic individuals displayed in I and J are not representative.
Number, integrity and generative transmission of integrated T-DNAs
Pollen produced by each of the four primary transgenic plants segregated with respect to gfp expression (Figure 1I). When embryos formed by the primary transgenic plants were germinated on a medium containing hygromycin, gfp expression was strong in both the scutellum and the emerging root, with a particularly high level in the root tip (Figure 1J). The selective medium discriminated between transgene carriers and non-carriers, with the former developing normally and the latter no longer able to grow after the emergence of the coleoptile, and unable to synthesize chlorophyll (Figure 2). The observed segregation ratio was consistent in three of the four cases with the presence of a single T-DNA insertion locus, while the progeny of the primary transgenic TG5E03 included significantly fewer transgene carriers than expected from a monogenic segregation (Table 2).
Figure 2. Rescue of segregating T1immature embryos from two of the four primary transgenics (TG5E01 and TG5E04). Seedling reaction to hygromycin was assessed after germination on hygromycin-containing K4N media for ten days. Hygromycin-resistant plantlets were considered as HPT positive (+), whereas susceptible ones were classified as HPT negative (−). HPT: hygromycin phosphotransferase (conferring hygromycin resistance).
Table 2. T1 segregation for hygromycin resistance
In order to comprehensively analyse the independent transgenic events obtained, T1 siblings were subjected to DNA gel blot analysis successively using HPT and gfp probes in order to characterize the integration sites. The gfp-profile of progeny of TG5E01 is given as an example in Figure 3. The HPT profiles shown in Figure 4A indicated that the number of T-DNA copies present in TG5E01, TG5E02, TG5E03 and TG5E04 was, respectively, two, one, two and two, while the gfp probe also highlighted a third copy in TG5E01 (Figure 3). The analysis also showed that all three T-DNA copies present in TG5E01 and both copies in TG5E04 segregated independently of one another, while the two copies in TG5E03 co-segregated (Figure 3 and 4A). A complete T-DNA has the size of 4762 bp. In the case of complete T-DNAs, the digestion of genomic DNA using HindIII is expected to result in hybridizing fragments larger than 3114 bp (Figure 4F), as was seen in all plants except the descendants from TG5E01, where the two fragments carrying copies #2 and #3 were smaller.
Figure 3. DNA gel blot analysis of TG5E01 T1segregants. Genomic DNA was digested by HindIII, gel separated and transferred onto a Hybond-N membrane and hybridized against a DIG labeled fragment of gfp. The copy numbers were designated according to Tables 3 and 4.
Figure 4. Analysis of transgene events. (A) DNA extracted from a set of T1 plants of each event subjected to DNA gel blotting, and probed with a DIG labeled fragment of HPT. Note that copy #3 of TG5E01 as revealed and indicated in Figure 3 was not highlighted by the HPT probe used here. (B) Detached leaf hygromycin assay 10 days after transfer to hygromycin-containing medium. (C) Amplification of the HPT coding region and the CaMV 35S terminator. (D) Amplification of the gfp coding region and the CaMV 35S terminator. (E) Duplex PCR targeting the CaMV 35S and the UBI-1 promoter, as well as the HPT and gfp coding sequence. (F) The T-DNA based transformation cassette. LB – left border, P35S – promoter of the CaMV 35S promoter, HPT – HYGROMYCIN PHOSPHOTRANSFERASE gene, T35S – terminator of the CaMV 35S coding sequence. TNOS – terminator of the NOPALINE SYNTHASE gene, gfp – synthetic green fluorescent protein gene (S65T), PUBI-int – maize UBI-1 promoter with first intron, RB – right border. The full length of the T-DNA, the relative position of the HindIII restriction site as well as the positions of probes used in A (HPT probe) and Figure 3 (gfp probe) as well as amplicons shown in C, D and E are indicated.
The derived structure of the transgenes in TG5E02 and TG5E03 was confirmed by PCR assays based on primer pairs spanning various parts of the two transgenes (Figure 4C-F); however, for TG5E01, the PCR analysis of T1 plant 6 carrying only the T-DNA copy #3 (see Figure 3, plant lane 6) suggested that the HPT expression cassette was absent (Figure 4C,D,E, plant 6). Likewise, T-DNA copy #2, which was present in TG5E01 T1 plant 19 in addition to copy #3, did not show amplicons in Figure 4C and upper E either, indicating that it was also truncated with respect to the HPT cassette. Another example of incomplete T-DNA was found in TG5E04, where the single copy present in T1 plant 3 was truncated concerning the HPT expression cassette as revealed by PCR analysis (Figure 4E, upper band). Moreover, the also missing lower band in Figure 4E suggests another deletion in the promoter-gfp junction, whereas the DNA gel blot analysis had demonstrated the presence of both the HPT and the gfp sequences in this plant, and the internal HPT and gfp sequences were both successfully amplified.
Transgene expression in the T1 and T2 generation
When T1 plant leaf segments were challenged with hygromycin, those carrying a functional copy of HPT remained green, whereas those lacking the transgene or a functional version became bleached (Figure 4B). In all cases, the outcome of the test concurred with the conclusions drawn from the PCR and/or DNA gel blot assays (Figure 4A-E). Note the intermediate reaction of the non-transformed control (Figure 4B), which suggested that this assay on its own is insufficiently diagnostic of the incorporation of a HPT transgene.
GFP was detectable in the root, leaf and mature caryopsis of plants carrying the single copy transgene (TG5E02) (Figure 5A-H). In particular, the GFP signal was largely confined to cytosol. The level of gfp expression varied considerably between cell types. In the leaf, it was highest in the chlorenchyma and the stomatal guard cells (Figure 5A,E). In the mature caryopsis, the most intense signal was present in the aleurone layer (Figure 5C, G) and in the nucellar projection (Figure 5D, H).
Figure 5. Localization of GFP accumulation using confocal laser scanning microscopy. Chlorenchyma cells and stomatal cells in the lower leaf epidermis of (A) non-transformed and (E) transgenic plants. (B) Non-transformed and (F) transgenic root tip. Transverse section through a mature (C) non-transformed and (G) transformed caryopsis, the latter showing a low level of GFP activity in the endosperm and a high level in the aleurone layer (arrow). Detail of a transverse section through a mature grain showing the nucellar projection in a (D) non-transformed and (H) transgenic plant. Very strong GFP activity was observed at the base and the lateral periphery of the nucellar projection. Chlorophyll or grain husk autofluorescence shown in red. Bars indicate 50 μm.
Twelve T2 populations and control descendants from an azygous T1 segregant were tested for the expression of gfp in the root tip (Table 3). As expected, some of these families segregated (TG5E01-12, TG5E03-1, TG5E03-7 and TG5E04-2), but it was possible to identify five non-segregating (presumably transgene-homozygous) plants. All TG5E01-6 T2 individuals accumulated GFP (Table 3, Figure 6) although they only contain a truncated T-DNA fragment (for DNA gel blot see Figure 3 plant lane 6, for T-DNA truncation see Figure 4C-F). In this case, T-DNA truncation along with independent segregation of multiple T-DNAs unintendedly resulted in a selectable marker-free segregant that is homozygous with regard to the gfp gene. By contrast, none of the TG5E04-3 T2 individuals tested expressed gfp, thereby confirming the missing PCR-fragment of the gfp expression cassette from the respective T1 plant as shown in Figure 4E (lower band). Given the outcome of the leaf hygromycin test and the missing PCR fragments in the T1 plant TG5E04-3 (Figure 4B,E and F), the conclusion drawn was that the transgene copy present in this plant comprised non-functional sequences with respect to both transgenes. All data regarding the integrity, functionality and segregation of the T-DNA copies are summarized in Table 4. With regard to the gene-of-interest (gfp), five out of eight independent transgene copies proved functional, while the presumed functionality of two further copies could not unequivocally be shown due to the presence of other T-DNAs. In all those cases, the observed phenotype could be explained by genotypic data of the plants.
Table 3. Transgene segregation in twelve T2 populations, based on gfp expression in seedling tissue
Figure 6. Reporter gene expression in T2grains after seven days of germination. (A) Illumination by white light and (B) excitation with far blue light; transgenic (TG5E01-6) and wild type (WT) control grains are indicated.
Table 4. The integrity, functionality and segregation of the T-DNA copies present in four primary transgenics
Our initial aim was to develop a robust protocol for the stable genetic transformation of winter triticale. The approach taken rested heavily on transformation protocols established for its parental species, wheat and rye. The amenability of immature embryos to co-cultivation with A. tumefaciens in liquid culture was trialed, as this provides an efficient means of processing immature embryos in barley [14], and the same approach is effective in rye [15]. Unfortunately, however, it does not seem to work well in wheat (unpublished data), where desiccation of the immature embryos appears to improve the transformation efficiency [16]. In practice, triticale behaved like wheat in this respect, as transformation was only obtained when the immature embryos were co-cultivated on filter paper soaked with co-culture medium (Table 1). A similar study focusing on the spring triticale cultivar 'Wanad' compared the effectiveness of the three selectable marker genes BAR, HPT and NPTII driven by one of maize UBI-1, cauliflower mosaic virus 35S or A. tumefaciens NOS promoter, respectively [12], and concluded that the best combination was NOS::NPTII, even though NOS performs poorly in a monocotyledonous host [17]. In the present study, the HPT selectable marker gene was preferred, a gene which has also proven useful e.g. in barley [18,19], wheat and maize [13]. The gfp reporter gene was an efficient tool for monitoring transgenesis and the subsequent expression of the transgene (Figure 1F-J) [13,20].
Two of the three multiple T-DNA insertion events involved independent integration sites. In barley, >50% of multiple transgenic events induced by agro-infection involved only one integration site [21]. A more comprehensive analysis of transformation outcomes has been made in Arabidopsis thaliana, where the number of T-DNA copies integrated at a single site appears to be dependent not only on the identity of the A. tumefaciens strain and the explant, but also on the transformation methodology as well as the origin of replication of the vector providing the T-DNA [22]. The occurrence of transformation events with multiple T-DNA copies being integrated in independent genomic loci of triticale opens up the opportunity to generate transgenic segregants with reduced copy number. Moreover, co-introduction of effector and selectable marker gene using two different T-DNAs may give rise to selectable marker-free transgenics after independent segregation of the loci in the T1. While a similar case unintendedly occurred in the present study (TG5E01 T1 plant 6), a directed approach using barley has recently been presented by Kapusi et al. [23]. Although Agrobacterium-mediated transformation generally results in the less frequent integration of truncated transgenes than biolistic transfer, as many as 44% of primary wheat transgenics have been shown to carry incomplete T-DNAs [24] with many involving truncations at the left T-DNA border [25]. In barley, meanwhile, only 3% (of 260 primary transgenics analysed) retained the full T-DNA [26]. Truncation of the T-DNA can be expected to result in a loss of transgene function, as was indeed the case in the present experiments that revealed truncations in 37.5% of the integrated T-DNAs analysed.
The non-Mendelian segregation of transgenes among T1 progeny is a commonplace observation, and several hypotheses have been promoted to explain this phenomenon, such as T0 chimerism, multiple independently assorting insertion loci and transgene silencing induced by multiple transgene copies or DNA rearrangements [24,27-29]. In some cases, false positives can arise due to the expression of non-incorporated transgene cassettes including those carried by persisting Agrobacterium[30]. Non-Mendelian transgene segregation has been noted in triticale [12], but since this observation was based on a histochemical reporter gene assay and did not include any DNA analysis, its basis could not be ascertained. In the present study, fewer transgenic progeny was obtained than expected in the case of TG5E03, which suggests this plant to be chimeric with regard to transgenicity. This interpretation is corroborated by DNA gel blot, PCR and leaf assay, which indicated that all functional elements be present in at least one of the two coupled T-DNA copies (Figure 4A-F). Nonetheless, one of the copies may have produced aberrant mRNA causing post-transcriptional gene silencing. However, the non-Mendelian segregation observed in the T2 families derived from TG5E03 is anticipated to be caused solely by transgene silencing in some siblings, because chimerism can generally be ruled out in generations later than T0.
As monitored using confocal laser scanning microscopy, gfp expression was widely distributed, but concentrated in the cytosol (Figure 5). This localization mirrors what has been observed in transgenic barley and wheat [31], where the level of reporter gene expression in the aleurone and the endosperm was comparable to that driven by either the barley bi-functional α-AMYLASE/SUBTILISIN INHIBITOR (ISA) or the wheat EARLY-MATURING (EM) promoter [31,32].
The transformation method used in the present study is a valuable tool for the genetic engineering of triticale. The availability of reliable transformation technology should encourage the application of current functional genomics technologies to triticale, and accelerate the biotechnological-based approach to its improvement. In the present study, we show that comprehensive molecular analyses are required for the correct interpretation of phenotypic data collected from transgenic plants.
Plant material
Grains of the winter triticale (x Triticosecale Wittmack) cultivar 'Bogo' were germinated at 14/12°C day/night under a 12 h photoperiod with a photon flux density during the light period of 136 μmol s-1 m-2. After three weeks, the seedlings were vernalized by cultivating for eight weeks at 4°C under an 8 h photoperiod, re-potted and then grown in a glasshouse (18/16°C day/night, 16 h photoperiod, 170 μmol s-1 m-2 photon flux density).
Choice of A. tumefaciens strain
The hypervirulent A. tumefaciens strain AGL-1 was initially used for infection. The strain harbors the binary vector pYF133 [33] which provides CaMV d35S::HPT as a selectable marker gene, a synthetic gfp sequence [34] driven by the maize UBI-1 promoter [35] as the reporter gene, and the pCAMBIA vector backbone [36] (Figure 7). The T-DNA borders are derived from a nopaline-producing Ti plasmid. Later experiments used a hypervirulent derivative of strain LBA4404 [37], which harbors the binary vector pSB187; this differs from pYF133 in the promoter chosen to drive HPT (CaMV 35S) and the vector backbone which was derived from pLH6000 [38] (Figure 7). Both pYF133 and pSB187 were introduced into A. tumefaciens via electroporation.
Figure 7. Schematic representation of T-DNA regions of the binary vectors used for transformation. LB - left border; Pd35S - CaMV double 35S promoter; P35S - CaMV 35S promoter; HPT - HYGROMYCIN PHOSPHOTRANSFERASE selectable marker gene conferring resistance to hygromycin; T35S - CaMV 35S terminator; TNOS - Agrobacterium NOPALINE SYNTHASE terminator; gfp - green fluorescent protein gene; PUBI-1 - maize UBIQUITIN-1 promoter; RB - right border.
Isolation of immature embryos and their pre-culture and co-cultivation with A. tumefaciens
Immature caryopses were harvested 12–16 days post pollination, bathed for 3 min in 70% ethanol, further disinfected by immersion in 5% w/v sodium hypochlorite, 0.1% v/v Tween 20 for 15 min and washed five times in sterile, distilled water. Immature embryos were excised and plated scutellum side up on pre-cultivation medium as described elsewhere [13]. A frozen glycerol stock of A. tumefaciens (200 μL sampled from a culture with an OD600 of 2.0 suspended in 200 μL 15% v/v glycerol) was seeded into 10 mL antibiotic-free CPY medium and cultured overnight at 28°C with shaking. Prior to inoculation, the pre-cultivated immature embryos were transferred into liquid co-cultivation medium (BCCM) [13], which was then immediately replaced by 600 μL A. tumefaciens culture. After vacuum infiltration (1 min at 500 mbar), the immature embryos were washed with BCCM. The pre-cultured and inoculated immature embryos were then either stacked on filter paper disks moistened with 300 μL BCCM or WCCM or incubated in 2.5 mL of the same media. The co-cultivation of explants and Agrobacterium took place in the dark at 21°C for 48–72 h without agitation. A total of 50 immature embryos divided into 5 sets comprising 10 embryos each were used per treatment.
Tissue culture
After co-cultivation, the immature embryos were transferred to BCIM [13] containing 150 mg·L-1 timentin to kill any remaining A. tumefaciens. Calli formed over the following two weeks were transferred to BCIM containing 25 mg L-1 hygromycin and 150 mg L-1 timentin for a further two weeks and then removed to K4N regeneration medium [39] containing 25 mg L-1 hygromycin and 150 mg L-1 timentin. The cultivation plates were exposed to 136 μmol s-1 m-2 photon flux density over a 16 h photoperiod. Regenerating shoots were transferred to glass tubes containing the same medium until the plantlets were vigorous enough to be potted into soil.
DNA analysis
The PCR template comprised genomic DNA extracted from ~100 mg snap-frozen leaf using DNAzol (Invitrogen, Karlsruhe, Germany), according to the manufacturer's instructions. PCRs were based on the amplification of 100 ng template primed by the sequences listed in Table 5. Amplicons were separated by agarose gel electrophoresis and visualized by Ethidium bromide staining (Figure 4). Plants testing positive with the PCR assays were subjected to DNA gel blot analysis to characterize the integration site(s) and transgene copy number. At least 25 μg genomic DNA, extracted as described in [40], was digested with HindIII, separated by agarose gel electrophoresis and blotted onto a Hybond N membrane. A gene-specific probe (gfp or HPT) was labeled with DIG as recommended by the supplier (Roche, Mannheim, Germany).
Table 5. Primer sequences used for the PCR analysis of putative transgenic regenerants
Embryo rescue and segregation analysis of hygromycin resistance
T1 caryopses were harvested 21–28 days after pollination and the immature embryos were placed for 24 h on Gamborg's B5 medium [41] to induce germination. They were then transferred on to K4N medium [39] supplemented with 100 mg L-1 hygromycin, and held for up to ten days at 24°C under a 16 h photoperiod.
Hygromycin leaf assay
HPT expression in leaf segments of putative transgenic plants was analyzed following [42], attaching the leaf segments by their base to MS culture medium [43] containing 200 mg L-1 hygromycin and 0.05% v/v Tween 20. The plates were incubated for 7–10 days at 24°C under a 16 h photoperiod.
Microscopic analyses
Florets were harvested prior to anthesis and pollen grains released into 0.4 M mannitol. The expression of gfp was analysed using an Axiovert 200M fluorescence microscope equipped with the 38HE filter set (Zeiss, Oberkochen, Germany). Leaf, root and mature grain samples were analyzed with a LSM 510 META confocal laser scanning microscope (Zeiss, Jena, Germany) using a 488 nm laser line for excitation. GFP signals were detected with a 505–530 nm band pass filter. Autofluorescence was recorded with a 650 nm long-pass filter.
Competing interests
The author(s) declare that they have no competing interests.
Authors' contributions
GH designed the transformation experiments, participated in the design and analysis of the molecular studies and drafted the manuscript. SO performed the transformation experiments and participated in drafting the manuscript. DD performed the microscopical studies. JZ and MM participated in the design and organisation of the study and helped to draft the manuscript. JK conceived of the study, and participated in its design and organisation and finalized the manuscript. All authors read and approved the final manuscript.
The authors thank Cornelia Marthe for her excellent technical assistance. We also thank Dr. Sylvia Broeders for providing the vector pSB187.
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Plant J 1994, 6:271-282. PubMed Abstract | Publisher Full Text
Ishida Y, Saito H, Ohta S, Hiei Y, Komari T, Kumashiro T: High efficiency transformation of maize (Zea mays L.) mediated by Agrobacterium tumefaciens.
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Langridge P, Brettschneider R, Lazzeri P, Lorz H: Transformation of cereals via Agrobacterium and the pollen pathway – a critical assessment.
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Furtado A, Henry RJ, Pellegrineschi A: Analysis of promoters in transgenic barley and wheat.
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Furtado A, Henry RJ: The wheat Em promoter drives reporter gene expression in embryo and aleurone tissue of transgenic barley and rice.
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Fang YD, Akula C, Altpeter F: Agrobacterium-mediated barley (Hordeum vulgare L.) transformation using green fluorescent protein as a visual marker and sequence analysis of T-DNA::barley genomic DNA junctions.
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Komari T, Hiei Y, Saito Y, Murai N, Kumashiro T: Vectors carrying two separate T-DNAs for co-transformation of higher plants mediated by Agrobacterium tumefaciens and segregation of transformants free from selection markers.
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Hausmann L, Töpfer R: Entwicklung von Plasmid-Vektoren.
Kumlehn J, Serazetdinova L, Hensel G, Becker D, Loerz H: Genetic transformation of barley (Hordeum vulgare L.) via infection of androgenetic pollen cultures with Agrobacterium tumefaciens.
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Pallotta MA, Graham RD, Langridge P, Sparrow DHB, Barker SJ: RFLP mapping of manganese efficiency in barley.
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What is Address Search?
Spokeo reverse address search helps people connect with their neighbors and find people by address. Search your neighborhood today to find out current residents, past residents, and family information. Find out who lives on Pennington Rd today.
Other Spellings
Pennington Rd is also spelled Pennington Road.
Walton, KY
Time Zone & Current Time
Walton, KY lies within the Eastern Standard Time (or EST). The current time in Eastern Standard Time is 08:37 PM on 11/25/2015.
Walton, KY
Top Streets
Agarwood Dr, Albright Ct, Alta Vista Dr, Andrews Rd, Appaloosa Dr, Arbor Run Dr, Ashwood Dr, Aylor Ln, Banklick Rd, and Beatrice Ave
Walton, KY
Population Demographics
Household Income
Home Value
33 years old
50% male
Local Ethnicity
Ethnicity Ratio
About Walton, KY
Walton is a city in Boone and Kenton Counties in the U.S. state of Kentucky. The population was 3,635 in the 2010 Census. (more on Wikipedia)


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I may mat mine - then again, I may not. That all depends on the image and what it looks like.
But one thing is certain: They will be unmounted. Not only because I don't have a mounting press (but my local framer has one, and does exellent work), but because my experience is that mounted prints have a significantly higher risk of being damaged in the mail. The mat is stiffer, but it will break instead of flexing. If the print is mounted, it will break with the mount. If it's only attached with hinge tape or photo corners it will be undamaged even if the mount is a total write-off!


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Compare BOB Revolution SE Black Jogger Single Seat Stroller to Baby City Select Amethyst Jogger Single Seat Stroller
Compare More Products
BOB Revolution SE Orange Jogger... $275
Add to comparisonBOB Ironman Navy Jogger Single Seat... $336
Add to comparisonMaclaren Volo Scarlet Umbrella... $149
Add to comparisonBritax Römer B-Ready Black Travel... $399
Add to comparisonBOB Revolution SE Duallie Navy... $536
Add to comparisonBOB Revolution SE Duallie Black... $536
Add to comparisonBaby City Mini Stone/Black Jogger... $249
Add to comparisonBOB Revolution SE Navy Jogger... $328
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300+ Rentals · Chapel Hill
300+ Rentals · Chapel Hill
This separate-entrance apartment in our North Forest Hills home contains a queen bed and a queen futon, a full bath, and a small kitchen. You'll be steps away from a charming lake, and six minutes by car to the corner of Franklin and Columbia!
Our studio apartment has its own private entrance, and includes a small, functional kitchen and bathroom as well a spacious living area. A queen-sized bed and a queen-sized futon make it workable for up to four guests. The apartment is all yours, and yours alone, for the length of your stay. We're also happy to welcome you to join us in the rest of our house when we're home. We are happy to interact with guests as much or as little as they choose. If you'd like to hang out in the main par...
Chapel Hill
Enjoy the peaceful view from the private deck; hike the Botanical Garden trails which start from our driveway. Also an easy walk to the Dean Dome, Kenan Stadium and UNC Hospitals. Big windows make it easy to watch deer in this quiet neighborhood.
Full kitchen and your own washer and dryer, iron and ironing board. You can enjoy your meals on your private covered deck. Covered parking space. The 800 sq. ft apartment includes a large family room, bedroom, bathroom and kitchen. Plenty of games on the bookcase. Separate entrance, just go down the steps to the right of the house (as you face it). We live upstairs. Lock box for easy check in if we're not home. Information packet provided, but hosts are also available for recommendatio...
Chapel Hill
This is a 10' x 10' bedroom that contains a single bed, window, desk, a bookcase, 4' mirror and a closet. There is a shared bathroom across the hall that is also used by several long term guests, but w/ 5-10' showers, it has not been an issue.
Room is clean and tidy w/ ceiling fan, central a/c & hardwood floors and part of a 2400 sq' single family home located in quiet neighborhood, on D and CL bus line, and within easy access to U.NC and Duke U., Whole Foods, East gate shopping ctr and University mall. Have 1 very friendly dog--Schatzien. Share bath and kitchen. Continental breakfast available. We live simply and value environmental sensitivity--reduce, reuse,& recycle everything possible, thermostat is set at 78 degrees in su...
Chapel Hill
Our 1 bed/1 bath apt in a quiet, wooded residential neighborhood, is just 3 miles to UNC & perfect for those visiting Chapel Hill, Carrboro & surrounding areas. Enjoy private exterior entrance, free off-street parking & a short walk to the bus line.
Our cozy 800 sq ft apartment is in a wooded, quiet residential neighborhood. It has a queen size bed, full bath, built-in kitchen with two-burner electric stove, full refrigerator, microwave and service for 4. The apartment has a private exterior entrance, but is attached to our house, with one shared wall. The whole house, including the apartment, are smoke and pet-free. Guests have access to the following amenities: - clean, 100% cotton sheets and towels - two closets - Wi-Fi - AC...
Chapel Hill
We live a block away from Franklin Street bars and restaurants and a 10 minute walk to the Old Well or the Carolina Inn. The room has a full bed, which comfortably sleeps two people. Shared clean bathroom. Wifi, no cable.
Near the heart of downtown Chapel Hill, walkable to Carrboro, eclectic interior with Americana, modern, and antique furnishings and art. An ever growing collection of vintage records to satiate a wide variety of tastes. Guests will have access to a full kitchen, plenty of coffee, a parking space, and a shared bathroom. Guests are welcome to use the space as they like and interact as little or as much as they would like. My boyfriend and I are always available for recommendations and ofte...
Chapel Hill
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Charming home in Historic district with a downstairs with 2 bedrooms, and a bath in between. Separate entrance. Located just 3 blocks from UNC Chapel Hill. Can use the kitchen and frig (upstairs). Relaxed atmosphere. Come and go freely.
Very private and quiet, yet minutes from Franklin Street and walking distance to UNC. We have a separate entrance you will be using. Your room(s) are downstairs and we live upstairs. You are welcome to use the kitchen and frig. We also have a lovely balcony that overlooks the trees, where you can relax. Recommendations for local places to visit, eat, and drive. Happy to help! Hosts have traveled the world and lived overseas. We are directly down the street from the UNC quad, and Fran...
Chapel Hill
This luxury apartment is spacious, full of natural light, and beautifully appointed. Walk to dinner, catch a UNC game, or enjoy a show at one of the local theaters. Put your feet up and enjoy the handpicked amenities. Relax. You deserve it.
This two bedroom, two-bathroom top-floor condo brings all of the best of city life to the country. Think granite countertops meet stainless steel appliances meets walk-in showers meets relaxation. Floor-to-ceiling windows let in tons of natural space, and the 10-foot ceilings make this 1200+ square foot condo feel roomy and spacious. ENTERTAINMENT The 55" mounted 4K TV (no cords!) has basic cable and Apple TV (think Netflix, Hulu Plus, HBO Go, Showtime Anytime, etc.) and is hooked up to a ...
Chapel Hill
The West Wing is guest quarters to a residence on 15 acres 5 miles south of Chapel Hill. It's an easy 10 minutes (5 miles) to downtown Chapel Hill, UNC, UNC Hospitals and Carrboro. Comfortable, clean, attractive, and secluded.
You will feel like you are in the middle of nowhere, a little rustic, a little retro, but you have easy access to Chapel Hill, UNC, UNC Hospitals, and Carrboro. The bed is a double (full) thus "cozy" for two. The kitchen has a microwave and refrigerator and most anything you might need for a short stay, but cooktop or oven. You can wander my 15 acres. Check out he orchard, the garden, the bees and the beautiful hardwoods. The grocery is a mile away and town only 4 miles. If you need t...
Chapel Hill
A separate entrance leads to the office, eat-in kitchen, and living room with fireplace and Murphy bed. A screened porch opens onto the private backyard with hammock. Guests are welcome to share meals and facilities at the Center for Human Science.
A separate entrance leads to the office, eat-in kitchen, and living room with fireplace and Murphy bed. A screened porch opens onto the private backyard with hammock. Guests are welcome to share meals and facilities at the Center for Human Science. (2 left) … Located with its own separate entrance at the back of the Pink House, this apartment is right in the heart of things yet extremely private. Short of living in a sorority, there is no closer place to the UNC Hospital or Schools of Medic...
Chapel Hill
Look for the lightning bolt for faster booking
This room is located on the 2nd floor of the brick house. Centrally located a few blocks away from UNC and hospitals area. It also shares a bathroom with another room. Ideal for families that need connecting rooms. Plenty of natural light and shelves.
Chapel Hill
Look for the lightning bolt for faster booking
Gorgeous, light filled studio in downtown Carrboro/Chapel Hill. Walking distance from the best restaurants, galleries, music venues, shops and bars the area has to offer as well as UNC campus.
Chapel Hill
NEW! NOW IT'S A KING-SIZED BED! This large, post-and-beam room over our garage is light and airy with your own deck and private entrance. (Not handicapped accessible.) Ours is a small, quiet, wooded neighborhood 2.5 miles from the University of North Carolina and the UNC Medical Center, as well as the many wonderful restaurants and exquisite beauty of Chapel Hill and Durham (14 miles east). [Raleigh (30 mi.)/RTP (15 mi) and RDU Airport (20 mi.) are within easy driving distance as well.]...
Chapel Hill
My retreat in the woods is cozy, clean, peaceful and oh so comfy. Amenities include double bed in a spacious room, wifi, scrumptious full breakfast and a cuddly black lab. Enjoy a walk in the woods in a tranquil environment just minutes from UNC.
Welcome. This listing is fairly new (Sept 2015) but I've been a SuperHost for awhile and have other rooms that have all received 5 star reviews. Just recently decided to add this listing to the group. This space is super cozy and can accommodate 2-3 people (I add a super comfortable futon for the 3rd person). I live in the middle of a beautiful tranquil woods just 5 minutes from UNC. Clean, quiet, comfortable, calm. Breakfast is served at 8 and is typically European style - coffee or t...
Chapel Hill
Spacious bedroom and shared bath in a 3-bedroom house located in a wooded neighborhood near UNC and downtown Chapel Hill/Carrboro. Amenities: queen bed, access to kitchen, living room, deck, central A/C, parking, (free!) bus, and high-speed wifi.
You are welcome to use the house common areas: living room, kitchen, deck, etc. I will be staying in another room in the house or nearby to help if need be. I tend to be a hands-off host. That being said, if you'd like some tips for how to navigate the area and what to do, I am happy to provide some suggestions! The house is in a quiet, wooded neighborhood filled with families and graduate students. It's only a 10-15 minute bus ride to campus yet you are likely to encounter deer on a morn...
Chapel Hill
We live in a large brick house with a private parking and within 15 min to campus or 20-25 min walk to Franklin Street. There are 3 residents, two of us share the downstairs bathroom. It's an old house, but beautiful. Lovely people too!
Chapel Hill
Look for the lightning bolt for faster booking
Clean and quiet. Walking distance to UNC campus, Franklin Street and Main Street in Carrboro. (The Chapel Hill bus system is free!) This place will offer a home away from home for one traveler at a time.
A townhouse in a chic subdivision near downtown Chapel Hill. Street parking is limited. Three minutes walk to bus stops (free transit). Walking distance to the campus. Great restaurants nearby. Bolin Creek for jogging and strolling. Rose garden in the Community Park. You will have your own bedroom and bathroom (hair dryer, towels, shampoo, and soap). You are welcome to relax in the living room. Kitchen access is limited (no cooking privilege). Simple breakfast is provided. Other bedrooms ...
Chapel Hill
This snug brick cottage is adjacent to UNC and two blocks from downtown. It boasts vaulted ceilings, tile floors, panelled walls, two patios, and a large lawn. Guests are welcome to share meals and facilities at the Center for Human Science.
This 1925 detached brick carriage house was converted during the depression to a "Honeymoon Cottage" for married UNC students. Since then, it has been completely and lavishly remodeled into a compact but luxurious home under the highest oak tree in Orange County. ... There are two brick patios for sitting out and enjoying the sunshine or the shade at any hour of the day while looking out over the expansive lawn. A gas grill makes making dinner a pleasure on those occasions you want to pass ...
Chapel Hill
Charming bed/bath new construction apartment. Hardwood floors in common space, carpet in bedroom. Informal dining off the kitchen. Beautiful scenic wooded views out of your window. Location is walkable to all conveniences. 3 miles to UNC, Carborro.
You will be sharing my charming, scenic, comfortable, quiet apartment with myself and my girlfriend. It offers a modern floor plan, with an efficient use of space, and no clutter. There is a full size washer and dryer in the apartment as well. The community offers a free coffee and tea bar every day to guests and residents. There are picnic tables and BBQ grills throughout the property, available for guests and residence. The other amenities on the property (pool, deck, community room) ar...
Chapel Hill
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A tag from the "Senior Giving Tree" could help make the holidays a little brighter for residents of local assisted living facilities.
For the fourth consecutive year the tree has been set up in the Die Brucke building between Macy's shoe and juniors departments. The tree's tags have gift suggestions for seniors who might not otherwise receive a present during the holiday season.
The tree's been up since just before Thanksgiving, but a number of tags remain unclaimed, organizers say.
In a story last year on the movement, organizer Mindy Stonebraker of Walla Walla's Pioneer House said seniors often tend to be forgotten. "We really want people to know there are really a lot of seniors in our community who don't have anything, or not much of anything," she said at the time.
In an email on the Senior Giving Tree this year, Stonebraker said coordinators were overwhelmed by the generosity of the public last year.
Those who want to participate can pick up a tag. Dropoff locations for the gifts are Pioneer House, 1018 Whitman St.; Quail Run Retirement Community, 1701 Plaza Way; and Eagle Meadows Assisted Living, 550 E. Whitman Drive in College Place.
Suggested ideas tend to be blankets, sweaters, slippers, candy and other items. Gifts should be unwrapped but placed in gift bags with the tag attached. For more information call 522-4630.
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Frumpy Mom: Trying to predict my daughter's future puts me in a hairy spot
By MARLA JO FISHER
Don't you wish you had the tiniest little crystal ball so you could peek into the future? I don't need to know what the stock market will be doing next year, or what the winning Lotto numbers will be. Well, OK, I do, but that's not what's on my mind right now. I just wish I knew if my 16-year-old daughter w


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April 18, 2012
gosh she is awesome
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gosh she is awesome
What a fuckin incredible woman. Isn't it about time we saw her cumming her tits off though? Your turn Heather!
naja kurz & knackig eben :-)
Very Very NAUGHTY girl!


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In another, they sit smiling inside their Los Angeles apartment at a messy kitchen table.
The tone veered wildly between camp, melodrama, realism, and messy amalgamations of the three.
Twenty tired people crowded into every corner of the small, messy room.
Not messy custody battles or an ugly war of words with her ex.
Swing voters are messy, unpredictable, and dissatisfied with both political parties, which drives people like Ruy Teixeira crazy.
The nurseries overflow with messy pets—nor does Puck ever remember to feed his guinea-pig.
Thousands of other planets will gain the settlers that messy Row loses.
And if he's at all messy about it, I give you leave to roll him downstairs.
Sorry to sacrifice you, honey, but the other way is just too messy.
It was not easy; it was a hot, messy, scratchy, grunting job.


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You are in a confined arena with no other horses creating a noise or mischief,
When you are feeling comfortable on your horse,
On a nice sunny day,
When all is serene
Then close your eyes.
Try to pick up on the movement of the horse by the sense of feel through the seat and thighs.
With your eyes closed, try to sense the sequence of the
Moving legs of the horse.
You can do the same without closing the eyes by looking straight ahead, but if you can close the eyes then the senses of feel, touch and motion are highlighted.


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|Regions with significant populations
|• India • Pakistan
The Zamindara or Rautara, (combinations of words Rai and Tartar from village Hasanpur-Beenapur) is a distinct Muslim community found mainly in the Indian state of Uttar Pradesh.[1] Members of the Zamindara community who migrated to Pakistan after the independence in 1947, are settled in Karachi, Sindh. It is a community in India who practice Central Asian Turkic-Mongol Culture in their lifestyle. The Zamindara are a sub-group within the wider Shaikh, community. Zamindara in Urdu means landowner.
Zamindara speak a distinct dialect of Urdu among themselves that is similar to a central Asian Turkic language. They refer to it as Azmiat, but use Urdu with outsiders. This community is also referred to as "Azmi Community" or "Azmi Biradari" or "Azmi Qaum" and "Rautara Biradari". The group practices endogamy, strictly prohibiting marriage outside their community.
Zamindara usually have Shaikh, Rautara, or Khan, as their surnames.
They practice Khorasan culture in their daily life.
They manage their own education system in their Senior Secondary School and Arabic Seminary. They are from the Hanafi School of Thought, but the youths of this community adopted the Ahle Hadith School of Thought without anyone's influence.
History and origin[edit]
The sheikh (warrior clan) are the original inhabitants of Azamgarh. They embraced Islam when they came in contact with warrior clans of Central Asia such as Turks, Mongols and the Pathans of Afghanistan. The Zamindara are said to be descendants of Turk and Rajput caste and Pathans who migrated to the region from Khorasan. Many Zamindaras were substantial landowners, and reference is made to this community in Abu'l-Fazl's, 16th-century magnum opus, Ain-i-Akbari, as the landowners of pargana Nizamabad.[1]
By the early 19th century, owing to their industry and enterprise, Zamindara's had substantially increased their landed possession. In addition, many were educated and had obtained good position in the professions and government services in erstwhile British India.[1]
The Zamindara today are mainly concentrated in Azamgarh District, but are also found in substantial number in neighbouring districts of Mau and Jaunpur, and in certain pockets of Gorakhpur and Sultanpur.
Zamindari are essentially a community of landowners and prior to the abolition of the Zamindari system, held large tracts of lands.
The population of Zamindara community in Azamgarh and Jaunpur number approximately 2.5 million of which 1.2 million are Turks and Pashtun and 1.3 million Shaikh.
The villages founded by Shaikh include Sanjarpur (King Sanjar Shaikh) Amilo (khas), Saat Gao, Mangrawan Raipur, Kundanpur Kotila, Bindawal- Jairajpur, Bisham, Nandao, Anjan Shahed, Mahwara , Ashrafpur and Mande ,
The villages founded by Pathans include Mohammedpur (Ghazni), Fariha (Farah) khandwari, Daudpur, Sherwaa and Surahi.
The villages founded by Mongol include Muslim Patti, Manjeerpatti, Mirzapur, Hasanpur-Beenapara and Shahpur.
This page uses Creative Commons Licensed content from Wikipedia. A portion of the proceeds from advertising on Digplanet goes to supporting Wikipedia.


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Project Manager (885) Warren, New Jersey
Business Description:
Through proprietary global panels and research products and services, Lightspeed Research delivers valuable data to help businesses make informed decisions.
We have significantly grown since we were incorporated in May 2000. Our progress presents new and exciting opportunities – requiring creativity, new technologies, thought leadership, and operational excellence.
Lightspeed Research is part of the Kantar Group, one of the world's largest research, insight and consultancy networks and the research arm of WPP — the world leader in communications services. We have offices around the globe offering a wide range of career opportunities.
Job Summary:
The Project Manager will have responsibility over a number of projects ensuring each project is set-up efficiently, liaising with the client and account director over issues affecting set-up and fieldwork, tackling issues as they arise (or bringing them to the attention of the appropriate person), ensuring established processes are followed during the project and that required documentation is kept up to date. The Project Manager will report to a Client Operations Manager. This role would suit a motivated professional who enjoys fostering relationships and working as part of a team, but also embraces responsibility for their work as an individual.
Project Management (80%)
* Manage every aspect of online project management, including
* client liaising / servicing.
* Communicating with panel partners and outsource operations company to ensure field requirements are met.
* Providing advice on project and questionnaire requirements.
* Survey scheduling and set-up.
* Sample specification.
* Survey monitoring.
* Data checking and validation.
* Working to a budget and being mindful of costs.
* Keep account director and pricing team updated on status.
Other Support tasks (20%)
* Support may be required in the following areas:
* Data preparation.
* Panel care – handling queries from panel care staff.
* Understanding incentive fulfillment.
Minimum Qualifications
* Bachelors Degree with 1 to 2 years of market research experience preferred.
Additional Qualifications
* Excellent communication skills – specifically how to distil the issues, manage client expectations and communicate in a timely manner under pressure.
* Client management experience, displaying a solutions driven approach.
* Familiarity with technological resources including Excel, SPSS and aptitude to use web-based survey tools.
* The successful project manager will exhibit the ability to be proactive, solution-oriented and well-organized.
* A good attention to detail and the ability to multi-task are essential.
* Experience in online research is an advantage but is not essential.


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Hey everybody,
I'm planning a self-contained trip from Jackson, WY to Portland, OR from mid July to mid Aug (around 30 days total). I'll be following the Transam route up through Missoula. But from Missoula I'm trying to figure out the best route to Portland. I like the idea of following the Columbia river, so I was thinking of following the Lewis & Clark trail. However when I looked at the description of the two routes between Missoula and Clarkston, WA, on the ACA map I saw warnings of fairly poor riding conditions consisting of stretches of road with no shoulders and dirt/gravel roads. I also heard that the head winds can be pretty bad along the Columbia gorge.
Should I, instead, follow the Transam route down into Baker City, and either head north towards the Columbia river, or would you suggest that I skip the Columbia river and stay on the Transam toward the coast and then make my way to Portland?
Is there a 3rd alternative that someone can recommend?
I'll be camping around 80-85% of the time, so a route with nice inexpensive campsites would be nice.