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VIDEO: Mina Leslie-Wujastyk climbs Tetris, hard 8A+by Björn Pohl - UKC Oct/2012
This news story has been read 4,195 times
Mina Leslie Wujastyk is currently road tripping through the US of A. Right now she is in Colorado from where she has good memories as this is where she did Mind matters, 8A+, which used to be her hardest boulder problems to date.
That's right, it used to be, because a few days ago she managed to take down David Graham's powerful and technical compression problem Tetris in the Wild Basin area of the Rocky Mountain National Park.
The problem was originally given 8B, but now seem to be settling around hard 8A+.
Climb Magazine caught up with her and asked her about it:
Mina: Tetris is a fairly new problem and, as is often the case with new lines, there is some disparity regarding the grade. It was originally given 8B by David Graham with some subsequent climbers suggesting 8A+ and some agreeing with 8B. I don't have much experience at this grade, but in my opinion, based on the experience I do have, I would suggest hard 8A+ as opposed to 8B. I also climbed the problem with a different sequence to Dave Graham, perhaps I found a path of less resistance? This is definitely my hardest problem so far :).
How many women have climbed 8b+ and 8A+ within one month (Mina climbed Mecca at Raven Tor 6 Sept)?
Here is the video:
Mina Leslie-Wujastyk is sponsored by: Five Ten, Arc'teryx, BMC, Element Chiropratic Clinic, Beta Climbing Designs and Optimum Nutrition
Tonight's Friday Night Video follows on nicely from our recent article 'The Occasional Alpinist's Guide to the... Read more
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Mina Leslie-Wujastyk has redpointed 8c at , making its first female ascent. First climbed by Mark Leach in 1989, the... Read more
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Company Overview of Newknow Network, S.A.
Executive Profile
Juan Moran
Founder, Chairman of the Board and Chief Executive Officer, Newknow Network, S.A.
|Total Calculated Compensation
|This person is connected to 1 Board Members in 1 different organizations across 2 different industries.
See Board Relationships
Juan Moran is the Founder of Newknow Network, S.A. and serves as its Chairman and Chief Executive Officer. Prior to Newknow, Mr. Meta4 NV founded two successful start-ups (MBS and Meta4 NV). He served as Chairman and Chief Executive Officer of Meta4 from 1992 to 1999.
Corporate Headquarters
Calle Navaluenga, 2Phone: 34 91 639 90 00
Las Rozas, Madrid 28230
Fax: 34 91 638 71 59
Board Members Memberships
Founder, Chairman of the Board and Chief Executive Officer
EducationThere is no Education data available.
Other Affiliations
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The Strategic Framework 2014-2019
The Strategic Framework is a capstone to strategic planning at Queen's. Its overarching goal of the strategic framework is to strengthen Queen's vision as Canada's quintessential balanced academy, ensuring we remain a university recognized equally for its research excellence and its transformative student learning experience.
At the framework's core are four interconnected strategic drivers: the student learning experience; research prominence; financial sustainability; and internationalization.
Download Queen's University Strategic Framework: March 2014 (500 KB).
Progress Reports
Annual progress reports on the Queen's University Strategic Framework will be shared with the Board of Trustees, the Senate and the Queen's community. This Initial Report was presented to the Board of Trustees in September 2014.
Please note that some of the metrics referenced in the reports are rarely, if ever, updated more frequently than annually, and sometimes even less frequently. For example, the National Survey on Student Engagement, the leading measure of undergraduate student engagement, is administered every three years. As well, a three-year rolling average of our share of federal granting council funds is calculated every two years.
If you require this document in an alternate format, please inquire through firstname.lastname@example.com or contact the Department of Marketing.
Developed by Principal Daniel Woolf, Provost Alan Harrison and the Board of Trustees, the Framework outlines a multi-year stratgy and a set of metrics and milestones through which the University's progress can be evaluated. Its structure is based on the foundations laid by the Academic Plan, Strategic research plan, which weer both approved by the Senate in the 2011-12 academic year, as well as other planning initiative (that were ongoing at the time) such as a plan for internationalization and the renewal of the Campus Master Plan.


text:
|By Marketwired .
Article Rating:
|January 4, 2013 08:00 AM EST
SANTA BARBARA, CA -- (Marketwire) -- 01/04/13 -- D2 Technologies, the market leader in embedded IP communications software platforms, today announced that chief operating officer Doug Makishima will join other key industry leaders to discuss how smarter devices and ubiquitous high-speed mobile data access via 4G and WiFi fuel the evolution of richer IP-based communication services such as HD voice, instant messaging, video calling and content sharing.
Mobile operators are now successfully deploying Voice over LTE (VoLTE) and rich communications suite (RCS) services that provide advanced communications capabilities popularized by Over-The-Top (OTT) offerings such as Skype, WhatsApp, FaceTime and enterprise Unified Communications (UC) solutions, but with carrier quality, reliability and reach. Mobile subscribers now have more options than ever before as their appetite grows for advanced communication modes beyond traditional voice and SMS.
"As mobile operators migrate legacy voice and SMS services to LTE networks using IMS, there's an opportunity to simultaneously provide rich communications via RCS," noted Makishima. "D2's mCUE 4G IMS client software provides mobile device OEMs and IC vendors with optimized and interoperable VoLTE and RCS capabilities."
Doug Mohney, editor-in-chief of HD Voice News, will moderate the panel, "Richer Mobile Communications through Broadband: Learn why the Apple iPhone 5 supports HD voice, MetroPCS favors 'rich' communications services (RCS) over HD voice, and what RCS apps are available through joyn and OTT. Enabled by faster HSPA+ and LTE networks, the mobile industry is moving to HD voice and a new wave of interactive RCS."
Alan Percy, senior director of strategic marketing, NA, AudioCodes and Jeffrey Rodman, co-founder and chief evangelist, Polycom, will join Makishima. CES 2013 will be held January 7-10 at the Las Vegas Convention Center. The panel discussion will occur Monday, January 7 at 5:00pm.
About D2 Technologies
D2 is the recognized leader in VoIP and converged IP communications software for devices used in 4G and next-generation networks that deliver advanced communication services, such as VoIP over LTE (VoLTE) and IMS-based Rich Communications (RCS) video call. Manufacturers and service providers rely on D2 software to deliver carrier-grade IP communications across any network (LTE, WiMAX, cellular, WiFi, broadband, PSTN), service (voice, video, IM chat, SMS, presence/status, etc.) and system (carrier, IPPBX, UC, OTT, social network, etc.) for a broad range of fixed and mobile devices. From processing billions of VoIP minutes each month to adding enhanced IP communications for Android, learn how D2 software is converging communications at www.D2Tech.com or follow the company on Twitter @D2Tech.
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Doug Makishima
D2 Technologies
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Jeff Denenholz
VOXUS, Inc.
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As more and more data is generated from a variety of connected devices, the need to get insights from this data and predict future behavior and trends is increasingly essential for businesses. Real-time stream processing is needed in a variety of different industries such as Manufacturing, Oil and Gas, Automobile, Finance, Online Retail, Smart Grids, and Healthcare. Azure Stream Analytics is a fully managed distributed stream computation service that provides low latency, scalable processing of streaming data in the cloud with an enterprise grade SLA. It features built-in integration with Azur...
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The broad selection of hardware, the rapid evolution of operating systems and the time-to-market for mobile apps has been so rapid that new challenges for developers and engineers arise every day. Security, testing, hosting, and other metrics have to be considered through the process. In his session at Big Data Expo, Walter Maguire, Chief Field Technologist, HP Big Data Group, at Hewlett-Packard, will discuss the challenges faced by developers and a composite Big Data applications builder, focusing on how to help solve the problems that developers are continuously battling.
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"The Internet of Things transforms the way organizations leverage machine data and gain insights from it," noted Splunk's CTO Snehal Antani, as Splunk announced accelerated momentum in Industrial Data and the IoT. The trend is driven by Splunk's continued investment in its products and partner ecosystem as well as the creativity of customers and the flexibility to deploy Splunk IoT solutions as software, cloud services or in a hybrid environment. Customers are using Splunk® solutions to collect and correlate data from control systems, sensors, mobile devices and IT systems for a variety of Ind...
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Clearly the way forward is to move to cloud be it bare metal, VMs or containers. One aspect of the current public clouds that is slowing this cloud migration is cloud lock-in. Every cloud vendor is trying to make it very difficult to move out once a customer has chosen their cloud. In his session at 17th Cloud Expo, Naveen Nimmu, CEO of Clouber, Inc., will advocate that making the inter-cloud migration as simple as changing airlines would help the entire industry to quickly adopt the cloud without worrying about any lock-in fears. In fact by having standard APIs for IaaS would help PaaS expl...
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Electric Winch Wireless Remote Control System
You can control your winch wirelessly with this wireless remote control system for most types of 12volt winches. This wireless winch control delivers convenient power in
free shipping
(3 Reviews)
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Forecum 4f Water-resistant Wireless Remote Control Doorbell Set - White + Yellow
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6. Receiver sensibility: -103dbm
7.Operating temperature: -10 ~ +70'C
8. Size: 67 x 47 x 17mm
9. Weight: 75g including the remote controller
10. Remote control distance
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Z07-5 Universal Handheld Wireless Bluetooth Remote Control Shutter Monopod - Black
Color: Black Brand: N A Model: Z07-5 Material: Aluminum alloy + silicone + ABS Quantity: 1Set Mount Type: Monopod Other Features: Supports Android 3.0 or IOS 4.0 above sy
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Wireless Digital Remote Control Electrical Switch
Please refer to : http: dx.com p wireless-5-ways-remote-control-home-lamp-light-switch-59215 - Color: White
- 86 wall plate panel
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Wireless Infrared Shutter Remote Control for Nikon DSLR Camera - Dark Blue
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RF Wireless Remote Control AC Power Socket (AC 220V/500W)
Includes: Remote controlled switch socket Remote control
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right moving Remote Type Radio Control Remote Control Frequency 49 MHz Remote Control Range 15 meters Battery Capacity 100 mAh Battery Type Li-ion Charging Time 20 minutes
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Wireless PCI Computer / PC Lock w/ Remote Control - Grass Green
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OUDI AD-88X Wireless Slide Remote Control - Silver + Black
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ZnDiy-BRY FC-1 Handheld Wireless Remote Control Frequency Tester
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Three Ways Wireless Digital Remote Control Switch - Black + White + Multi-Colored
: 220V; Frequency: 315MHz; Load total power: 3000W; Remote control range: 20~50 meters; Working temperature: 85 centigrade; Widely used in home, company, hotel, lightning
Mele F10-Pro 2.4GHz Wireless Air Mouse + Keyboard + Remote Control + Voice Mic
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FORECUM FK-6 Wireless Digital Remote Control Door Bell - Black
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text:
Shout Out: Advertising
- In the New York area, there's a store called B&H, whose advertisements involve a guy named Dave and a robot named Hal.
- There's a Honda advertisement that uses the opening to the Waldstein Sonata by Beethoven.
- Some advertising for Sonic Heroes referenced The A-Team quite heavily, such as a magazine ad◊ aping the famous opening narration or the (now-defunct) website using a slightly changed snippet of the theme.
- Trojan Condoms: In The Iliad the Greek army gets into a giant horse, which gets through the enemy's defenses. Men inside a horse, inside the enemy's walls. Pure Faux Symbolism. And then, after penetrating the enemy fortress, the horse bursts open and all the soldiers inside start rampaging everywhere.
- Advertising billboards for a flatrate internet package included Doom HUD as seen here◊ and here.
- A Filipino snack ad best described, as one comment put it, as "SACRILEGE TO LUCKY STAR." In terms of character designs, Sailor Moon wasn't spared either.


text:
The last piece of creative we produced for the Florida Tobacco Pilot Program (Truth) in 2000.
Objective: Demonstrate to teens that their rebellious notion to smoke was actually secretly engineered by middle-aged, corporate white guys. Sure, faux movie trailers are now a well-worn ad device. But I think this is an effective head fake and I'm proud to say that this cinema spot came out precisely the way I wrote it (word for word) and was edited precisely the way Paul Keister and I boarded it (scene for scene). Directed by Phil Joanou. Edited by Jeff Sternberger and Paul Norling.
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text: Beautiful hotel and gorgeous pool area. The staff in the pool area were very helpful and accomodating. The beach access is just across the beach bike/walk path which takes you all the way up and down the beach. However, beware the Wifi Internet charge- ~ $15 per 24 hour period...little pricey for the already high priced rooms.


text:
Page 1
Molecular Biology of the Cell
Vol. 18, 1044–1055, March 2007
Increased Common Fragile Site Expression, Cell
Proliferation Defects, and Apoptosis following Conditional
Inactivation of Mouse Hus1 in Primary Cultured Cells
Min Zhu and Robert S. Weiss
Department of Biomedical Sciences, Cornell University, Ithaca, NY 14853
Submitted October 27, 2006; Revised December 21, 2006; Accepted December 29, 2006
Monitoring Editor: Orna Cohen-Fix
Targeted disruption of the mouse Hus1 cell cycle checkpoint gene results in embryonic lethality and proliferative arrest
in cultured cells. To investigate the essential functions of Hus1, we developed a system for the regulated inactivation of
mouse Hus1 in primary fibroblasts. Inactivation of a loxP site-flanked conditional Hus1 allele by using a cre-expressing
adenovirus resulted in reduced cell doubling, cell cycle alterations, and increased apoptosis. These phenotypes were
associated with a significantly increased frequency of gross chromosomal abnormalities and an S-phase–specific accu-
mulation of phosphorylated histone H2AX, an indicator of double-stranded DNA breaks. To determine whether these
chromosomal abnormalities occurred randomly or at specific genomic regions, we assessed the stability of common fragile
sites, chromosomal loci that are prone to breakage in cells undergoing replication stress. Hus1 was found to be essential
for fragile site stability, because spontaneous chromosomal abnormalities occurred preferentially at common fragile sites
upon conditional Hus1 inactivation. Although p53 levels increased after Hus1 loss, deletion of p53 failed to rescue the
cell-doubling defect or increased apoptosis in conditional Hus1 knockout cells. In summary, we propose that Hus1 loss
leads to chromosomal instability during DNA replication, triggering increased apoptosis and impaired proliferation
through p53-independent mechanisms.
Cell cycle checkpoints monitor the fidelity of chromosome
replication and segregation. In response to genome damage,
checkpoint signaling induces cell cycle arrest and promotes
DNA repair, or alternatively, it triggers apoptosis to elimi-
nate damaged cells. Mammalian DNA damage responses
are coordinated by two primary checkpoint pathways that
center on the phosphatidyl inositol kinase-like protein ki-
nases ataxia telangiectasia mutated (Atm) and Atm- and
Rad3-related (Atr) (Bakkenist and Kastan, 2004). An Atm-
dependent pathway responds to double-stranded DNA
breaks (DSBs) such as those caused by ionizing radiation,
whereas an Atr-dependent pathway is activated by a variety
of DNA lesions, including bulky DNA lesions and replica-
tion stress as well as DSBs.
Optimal Atr signaling requires its binding partner, Atrip, as
well as additional accessory factors TopBP1, Brca1, Claspin,
and the Rad9–Rad1–Hus1 (9-1-1) complex (Shechter et al.,
2004b). The 9-1-1 complex shares predicted structural similar-
ity with the sliding clamp proliferating cell nuclear antigen and
is loaded onto chromatin at damage sites by a clamp loader
Karnitz, 2003). 9-1-1 promotes the phosphorylation of Atr sub-
strates such as Chk1, Rad17, and Rad9 itself (Weiss et al., 2002;
Zou et al., 2002; Roos-Mattjus et al., 2003; Bao et al., 2004) and is
required for an intra-S cell cycle checkpoint that represses
DNA synthesis after DNA damage (Roos-Mattjus et al., 2003;
Weiss et al., 2003; Bao et al., 2004; Wang et al., 2004b). Addi-
tional evidence indicates that the 9-1-1 complex also has a
direct role in DNA repair. The 9-1-1 complex physically
associates with multiple translesion DNA polymerases (Kai
and Wang, 2003; Sabbioneda et al., 2005) as well as base
excision repair factors, including the MYH DNA glycosy-
lase, DNA polymerase ?, flap endonuclease I, and DNA
ligase I (Toueille et al., 2004; Wang et al., 2004a. 2006a; Chang
and Lu, 2005; Friedrich-Heineken et al., 2005; Smirnova et al.,
2005; Shi et al., 2006). The 9-1-1 complex additionally is
required for homologous recombinational repair (Pandita et
al., 2006; Wang et al., 2006b). Consistent with its important
roles in cell cycle control and DNA repair, impaired 9-1-1
function is associated with cellular hypersensitivity to rep-
lication inhibitors and DNA damaging agents (Weiss et al.,
2000, 2003; Kinzel et al., 2002; Roos-Mattjus et al., 2003;
Hopkins et al., 2004; Wang et al., 2004b, 2006b).
Targeted disruption of components of the Atr-dependent
checkpoint pathway in mice causes embryonic lethality. De-
letion of Atr or Chk1 results in peri-implantation lethality
(Brown and Baltimore, 2000; de Klein et al., 2000; Liu et al.,
2000; Takai et al., 2000), whereas inactivation of Hus1, Rad9,
or Rad17 causes midgestational embryonic lethality (Weiss et
al., 2000; Budzowska et al., 2004; Hopkins et al., 2004). The
essential nature of these genes highlights the critical, yet
poorly understood, function of this pathway during an un-
perturbed cell cycle. In the course of a normal cell cycle, the
9-1-1 complex and other checkpoint components can be
detected in association with chromatin (Guo et al., 2000;
Hekmat-Nejad et al., 2000; Roos-Mattjus et al., 2002; You et
al., 2002; Zou et al., 2002; Jiang et al., 2003; Lee et al., 2003;
Dart et al., 2004). Even in the absence of extrinsic stress,
checkpoint signaling inhibits the cell cycle phosphatases
Cdc25A and Cdc25B, regulates origin firing, and suppresses
This article was published online ahead of print in MBC in Press
on January 10, 2007.
Address correspondence to: Robert S. Weiss (email@example.com).
1044© 2007 by The American Society for Cell Biology
Page 2
premature entry into mitosis (Miao et al., 2003; Shechter et
al., 2004a; Sorensen et al., 2004; Niida et al., 2005; Syljuasen et
al., 2005; Schmitt et al., 2006).
The Atr-dependent checkpoint pathway is also thought to
play a critical role in stabilizing stalled replication forks and
promoting fork restart (Lopes et al., 2001; Tercero and Diff-
ley, 2001; Sogo et al., 2002; Trenz et al., 2006). Possibly due to
failure of these important processes, certain yeast checkpoint
mutants show defects in the elongation step of DNA repli-
cation and accumulate chromosomal breaks at particular,
nonrandom genomic regions (Cha and Kleckner, 2002;
Raveendranathan et al., 2006). These sites may be analogous
to vertebrate common fragile sites (CFSs), chromosomal re-
gions where gaps and breaks frequently arise in metaphase
chromosomes prepared from cells under conditions of rep-
lication stress. Recent studies indicate that several compo-
nents of the DNA damage checkpoint machinery, including
Atr (Casper et al., 2002), Chk1 (Durkin et al., 2006), Brca1
(Arlt et al., 2004), and TopBP1 (Kim et al., 2005), among
others, are essential for maintaining CFS stability. No pri-
mary sequence conservation has been identified at CFSs, but
generally these sites are relatively AT rich, highly flexible,
and late replicating (Glover et al., 2005). These properties
suggest that CFSs might be prone to form secondary structures
that inhibit the progression of replication forks, creating a
requirement for cell cycle delay and replication fork stabiliza-
tion or repair by the checkpoint machinery (Cimprich, 2003).
Understanding the molecular basis for fragile site stability has
important implications, because these regions are frequently
deleted or rearranged in cancer cells (Arlt et al., 2006).
Previous attempts at molecular analysis of the essential
functions of Hus1 by using a conventional gene targeting
approach were complicated by severe phenotypes, includ-
ing midgestational lethality in embryos and proliferative
arrest in mouse embryonic fibroblasts (MEFs) (Weiss et al.,
2000). Successful culturing of Hus1-deficient cells from a
constitutive knockout mouse model additionally required
deletion of the checkpoint genes p21 or p53 (Weiss et al.,
2000; our unpublished data). Furthermore, because embryos
lacking both Hus1 and either p21 or p53 remained under-
sized and developmentally delayed, sufficient numbers of
cells for experimental analysis could be obtained only with
immortalized cultures. In this report, we describe a system
for the regulated deletion of Hus1 in primary cultured cells,
for use in dissecting the immediate consequences of Hus1
inactivation. By infecting primary MEFs containing a loxP
site-flanked conditional Hus1 allele with a cre-expressing
recombinant adenovirus (Ad-cre), we generated and ana-
lyzed large populations of Hus1-deficient and control cells in
vitro. Our results indicate that Hus1 inactivation results in
impaired cell proliferation and apoptosis associated with
CFS expression and S-phase–specific DSB accumulation.
Mouse Strains and Cell Culture
Previously described Hus1floxand Hus1?1mice were maintained on an 129S6
inbred genetic background (Weiss et al., 2000; Levitt et al., 2005). p53?/?mice
harboring the Trp53tm1Tyjallele were maintained on a C57BL/6J background
(Jacks et al., 1994). Mice were housed in accordance with institutional animal
care and use guidelines. MEFs were prepared from 13.5 dpc embryos from
timed matings between Hus1flox/floxand Hus1?/?1mice or from Hus1flox/flox
p53?/?and Hus1?/?1p53?/?mice. Briefly, embryos were dissected from the
deciduum, mechanically disrupted, and cultured in DMEM supplemented
with 10% fetal bovine serum, 1.0 mM l-glutamine, 0.1 mM minimal essential
medium nonessential amino acids, 100 ?g/ml streptomycin sulfate, and 100
U/ml penicillin. The initial plating was defined as passage zero (p0). MEFs at
p1 or p2 were used for all experiments.
Viral Infections
Ad-cre, an adenovirus that expresses Cre from the cytomegalovirus promoter
(University of Iowa Gene Transfer Vector Core, Iowa City, IA) (Stec et al.,
1999), was prepared in 293 cells. Briefly, cells were harvested at 48–72 h
postinfection, and viral lysate was subjected to CsCl gradient ultracentrifu-
gation at 63,000 rpm at 14°C for 7 h. Virus was further purified with a PD-10
desalting column (GE Healthcare, Little Chalfont, Buckinghamshire, United
Kingdom), and virus titer was estimated by spectrophotometry according to
the formula: 1 OD280? 1012virus particles/ml. For infections, 1 ? 106MEFs
were plated into a 10-cm culture dish and grown for 1 d. Cells were infected
with 1.95 ? 1011Ad-cre particles in 2.5 ml of culture medium at 37°C for 6 h,
after which time the virus was removed and fresh medium was added. Unless
otherwise specified, cells were passaged at 1 d postinfection (dpi) and then
maintained on a 3T3 culture schedule in which 1 ? 106cells were passaged
onto a 10-cm culture dish every 3 d (Todaro and Green, 1963).
Southern and Northern Blotting
Genomic DNA for Southern blotting was isolated from MEFs by proteinase K
digestion and precipitation with ethanol. DNA was digested with NheI, run
through a 0.8% agarose gel, transferred to a nylon membrane, and hybridized
with a32P-labeled 190-base pair EagI fragment from plasmid pCR2.1-5?UTR-
?2,3 (Levitt et al., 2005). For Northern blotting, total RNA was prepared from
MEFs by using RNA STAT-60 reagent (Tel-Test, Friendswood, TX), and
poly(A)?mRNA was isolated with biotinylated oligo(dT) (Promega. Madi-
son, WI). Purified mRNA was resolved on a 1% agarose/formaldehyde gel,
transferred to a nylon membrane, and hybridized with a32P-labeled cDNA
probe containing the entire mouse Hus1 open reading frame as described
previously (Weiss et al., 1999). After stripping, the membrane was hybridized
to a32P-labeled mouse Gapdh cDNA probe.
Cell Proliferation Assays and Cell Cycle Analysis
For cell proliferation assays, triplicate cultures were maintained on a 3T3
culture schedule. Population doublings (PDLs) were calculated using the
formula ?PDL ? log(nf/n0)/log2, where n0is the initial number of cells and
nfis the final number of cells (Blasco et al., 1997). For cell cycle analysis, 1 ?
106cells were plated per 10-cm culture dish 24 h before analysis. The next day,
the cells were incubated with 10 ?M bromodeoxyuridine (BrdU) for 45 min,
harvested by trypsinization, washed once in phosphate-buffered saline (PBS),
and fixed in 70% ethanol at ?20°C. The cells were then incubated in 2 N HCl,
0.5% Triton X-100, washed twice with 0.1 M Na2B4O7?10H2O, pH 8.5, incu-
bated with fluorescein isothiocyanate (FITC)-conjugated anti-BrdU (BD Bio-
sciences, Franklin Lakes, NJ) for 30 min at room temperature (RT), washed,
treated with RNAse A, and stained with propidium iodide (PI). Flow cytom-
etry was performed on a FACScan flow cytometer (BD Biosciences).
Apoptosis Assays
Cells grown in 10-cm culture dishes were collected by trypsinization along
with floating cells in the culture medium, washed twice with PBS at 4°C, and
resuspended in 1? binding buffer (10 mM HEPES, pH 7.4, 140 mM NaCl, 2.5
mM CaCl2) at a concentration of 1 ? 106cells/ml. Cells (1 ? 105) were then
incubated with Annexin V-FITC (BD Biosciences) and PI for 15 min at RT.
Flow cytometry was performed on a FACScan flow cytometer (BD Bio-
sciences) within 1 h.
Indirect Immunofluorescence Assays (IFAs)
Cells grown on coverslips were fixed in 2% paraformaldehyde in TBS for 35
min at 4°C (for ?-H2AX IFA) or in methanol at ?20°C for 30 min followed by
ice-cold acetone for two seconds (for p53 IFA). Cells were then incubated in
3% bovine serum albumin (BSA), 0.01% skim milk, 0.2% Triton X-100 in
Tris-buffered saline (TBS) for 20 min at RT. For ?-H2AX IFA, cells were
incubated with primary anti-?-H2AX antibody (JBW301; Upstate Biotechnol-
ogy, Lake Placid, NY) at 1:500 for 45 min, followed by secondary goat
anti-mouse Ig (H?L)-FITC (Southern Biotechnology Associates, Birmingham,
AL) at 1:60 for 35 min. For p53 IFA, cells were incubated with primary
anti-p53 antibody (FL393; Santa Cruz Biotechnology, Santa Cruz, CA) at 1:60
dilution at RT for 1 h, followed by secondary goat anti-rabbit Ig (H?L)-FITC
(Southern Biotechnology Associates) at 1:60 at RT for 35 min. Cells were
counterstained with 33 ng/ml 4?,6-diamidino-2-phenylindole (DAPI) for 1
Fluorescence-activated Cell Sorting (FACS) Analysis of
Cells (1 ? 106) were fixed in ice-cold 70% ethanol, incubated with 1% BSA,
0.25% Triton X-100 in TBS for 15 min on ice, and stained with primary
anti-?-H2AX antibody (JBW301, Upstate Biotechnology) at 1:500 overnight at
4°C. The next day, the cells were stained with secondary goat anti-mouse Ig
(H?L)-FITC (Southern Biotechnology Associates) at 1:400 for 30 min at RT
and counterstained with 5 ?g/ml PI containing RNAse A for 30 min at RT.
Flow cytometry was performed on an LSR II flow cytometer (BD Biosciences).
Regulated Hus1 Deletion in Primary Cells
Vol. 18, March 20071045
Page 3
Metaphase Chromosome Preparation and Fluorescence In
Situ Hybridization (FISH)
Metaphase spreads were prepared as described previously (Weiss et al., 2000).
Briefly, cells were incubated in culture medium containing 0.15 ?g/ml col-
cemid for 1 h and then trypsinized, incubated in hypotonic buffer (0.05 M KCl,
0.0034 M trisodium citrate) for 12 min at 37°C, and fixed for at least 20 min on
ice in 75% methanol, 25% acetic acid. Cells were then spotted onto microscope
slides and stained with 2% Giemsa in Gurr buffer, pH 7.0. Metaphase chro-
mosomes were scored under a 100? oil objective lens according to standard
guidelines (Savage, 1976; Mitelman, 1995). For FISH, unstained metaphase
chromosomes on slides were denatured in 70% formamide/2? standard
saline citrate (SSC) at 70°C for 2 min. Bacterial artificial chromosomes (BACs)
containing mouse genomic sequence mapped to fragile site regions were used
as probes in FISH analysis. Probe BAC-CITB-57C24 (Open Biosystems, Hunts-
ville, AL) was used to detect mouse Fra8E1 (Krummel et al., 2002), and
BAC-CITB-316M9 and BAC-CITB-513J1 (Open Biosystems) were used to de-
tect mouse Fra6C1 (Rozier et al., 2004). Probes were labeled with Spec-
trumGreen-dUTP (Vysis, Downers Grove, IL) by nick translation, ethanol
precipitated in the presence of mouse Cot-1 DNA (Invitrogen, Carlsbad, CA),
resuspended in deionized formamide, and incubated in hybridization buffer
(20% dextran sulfate/2? SSC, pH 7.0) at 37°C for 10 min. Probes were then
denatured at 75°C for 10 min, incubated at 42°C for 30 min, and hybridized
with metaphase chromosomes at 37°C for 24 h. After hybridization, slides
were washed three times each in 50% formamide/2? SSC at 42°C, 2? SSC at
37°C, and 0.1? SSC at 60°C, followed by a single wash in 4? SSC/0.1%
Tween 20 at RT. Slides were then counterstained with 33 ng/ml DAPI for 1
min. FISH signal was examined using a DMRE fluorescence microscope
(Leica Microsystems, Deerfield, IL).
Rapid and Complete Deletion of Hus1 in Primary
Cultured Cells by Cre-mediated Recombination
To examine the effects of acute Hus1 loss in primary cultured
cells, we established a genetic system for the regulated inacti-
vation of Hus1 in MEFs. This system is based on a conditional
allele, Hus1flox, in which exons 2 and 3 are flanked by loxP sites,
recognition sequences for the cre recombinase (Figure 1A).
Previous studies indicated that cre-mediated recombination at
Hus1floxdeletes exons 2 and 3, producing a null allele, Hus1?2,3,
which has the capacity to encode only the first 19 of 281 Hus1
amino acids (Levitt et al., 2005). For conditional Hus1 inactiva-
a constitutive null allele in which exon 1 and the start codon
have been deleted (Weiss et al., 2000), or wild-type Hus1 as a
control, and then performed infections with Ad-cre. In both
Hus1flox/?and Hus1flox/?1MEFs, cre-mediated recombination
was predicted to convert the conditional allele into the null
allele Hus1?2,3. Ad-cre infected Hus1flox/?cells would continue
to express Hus1 from the remaining wild-type allele, whereas
Ad-cre–infected Hus1flox/?1cells would produce no functional
Hus1 transcripts (Figure 1A).
Initially, the minimal dose of Ad-cre required for com-
plete deletion of the Hus1floxallele was determined to min-
imize the cellular toxicity caused by cre (Loonstra et al., 2001;
Silver and Livingston, 2001). Hus1flox/?cells were infected
with various doses of Ad-cre, and genomic DNA was iso-
lated at 2 dpi and subjected to Southern blot analysis. Once
the minimal effective Ad-cre dose was determined (data not
shown), the kinetics of Hus1 inactivation were examined. By
1 d after infection with the minimal Ad-cre dose, the Hus1flox
allele was fully converted to Hus1?2,3in both Hus1flox/?and
Hus1flox/?1MEFs (Figure 1B). Northern blot analysis also was
performed to evaluate Hus1 expression at the corresponding
times after Ad-cre infection. As shown in Figure 1C, mock-
infected Hus1flox/?and Hus1flox/?1cells both expressed wild-
type Hus1 transcripts, with the expression level being higher
in Hus1flox/?cells than Hus1flox/?1cells as expected. As rap-
idly as 1 dpi, wild-type Hus1 transcripts were no longer
detectable in Ad-cre–infected Hus1flox/?1cells and were re-
placed with a lower-molecular-weight mRNA correspond-
ing to the nonfunctional Hus1?2,3transcript. Ad-cre–infected
Hus1flox/?cells expressed both wild-type Hus1 and Hus1?2,3
transcripts as anticipated. In short, these results establish
that Ad-cre infection of conditional knockout MEFs allows
for the rapid and efficient inactivation of Hus1.
Figure 1.
using Ad-cre. (A) Schematic of the system for conditional inactiva-
tion of Hus1. The various Hus1 alleles used are indicated, with the
first several exons shown. Hus1floxis a conditional Hus1 allele in
which exons 2 and 3 are flanked by loxP sites (black triangles). After
Ad-cre infection, exons 2 and 3 of the Hus1floxallele are deleted,
producing the null allele Hus1?2,3. A nonfunctional Hus1 transcript
lacking exons 2 and 3 is produced from Hus1?2,3and is represented
by a dotted line. Hus1?1is a constitutive null allele that lacks exon
1 and additional upstream sequences. Ad-cre–infected Hus1flox/?
MEFs continue to express wild-type Hus1 from Hus1?, whereas
Ad-cre–infected Hus1flox/?1MEFs fail to produce any functional
Hus1 transcripts. (B) Southern blot analysis of Hus1 deletion after
Ad-cre infection. Genomic DNA was prepared from Hus1flox/?and
Hus1flox/?1MEFs at 1, 2, 3, or 4 d postinfection with Ad-cre or at 2 d
after mock infection (U, uninfected) and then subjected to Southern
blot analysis. The cells used in this experiment were not passaged
after Ad-cre or mock infection The positions of Hus1flox, Hus1?, and
Hus1?2,3bands are indicated. The Hus1?1allele is not detected in
this assay. (C) Northern blot analysis of mRNA prepared from cells
prepared Ad-cre or mock-infected Hus1flox/?and Hus1flox/?1MEFs.
The positions of the transcripts produced from Hus1flox, Hus1?, and
Hus1?2,3are indicated.
Conditional inactivation of Hus1 in primary MEFs by
M. Zhu and R. S. Weiss
Molecular Biology of the Cell1046
Page 4
Hus1 Loss Results in Decreased Cell Proliferation and
Increased Apoptosis
Fibroblasts derived from Hus1-deficient embryos fail to pro-
liferate in culture (Weiss et al., 2000). However, these exper-
iments are complicated by the fact that the Hus1-null cells
must be obtained from morphologically abnormal embryos
at a much earlier developmental stage than is typical for
MEF culture. The conditional Hus1 knockout system offered
the opportunity to rapidly inactivate Hus1 in a large popu-
lation of normal MEFs and to test the immediate impact of
Hus1 deficiency in a controlled setting. To examine the effect
of Hus1 loss on cell proliferation, we first quantified PDLs
for mock-infected and Ad-cre–infected Hus1flox/?
Hus1flox/?1MEFs cultured on a conventional 3T3 passage
schedule. As shown in Figure 2A, MEFs of all genotypes
initially doubled similarly. However, after 4 dpi Ad-cre–
infected Hus1flox/?1MEFs accumulated significantly fewer
PDLs than control Ad-cre–infected Hus1flox/?MEFs or mock-
infected Hus1flox/?1MEFs. These results identify an impor-
tant role for Hus1 in cell doubling under normal growth
The reduced cell doubling in Hus1 conditional knockout
cells could be due to cell cycle arrest, apoptosis, or both. To
differentiate between these possibilities, we first monitored
the progression of Hus1flox/?and Hus1flox/?1MEFs through
the cell cycle after Ad-cre infection. Asynchronous cultures
were harvested after labeling with BrdU at 2, 4, or 6 dpi.
Analysis of cell cycle distribution by bivariate FACS re-
vealed no significant differences between cells of the various
Hus1 genotypes at 2 dpi (Figure 2, B and C). However, at
Figure 2.
Hus1flox/?and Hus1flox/?1MEFs at passage one were infectectd with Ad-cre or mock infected and then cultivated following a 3T3 culture
schedule as described in Materials and Methods. Plot shows the number of accumulated PDLs. (B) Schematic representation of a FACS dot plot
of cell cycle distribution. Staining intensity for PI (x-axis) is plotted versus that for anti-BrdU-FITC (y-axis). The S-phase population is divided
into S1 (BrdU positive) and S2 (BrdU negative). (C) Effects of Hus1 loss on cell cycle distribution. MEFs of the indicated genotypes were
labeled with BrdU at the indicated times postinfection or mock infection, stained with anti-BrdU and PI, and analyzed by flow cytometry.
Cells were passaged 1 d before BrdU labeling. The percentage of cells in each phase of the cell cycle is indicated. (D and E) Increased apoptosis
after conditional inactivation of Hus1. MEFs of the indicated genotypes were stained with Annexin V-FITC and PI at the indicated times
postinfection or mock infection and analyzed by flow cytometry. (D) Plot shows the percentage of apoptotic cells (Annexin V positive, PI
negative). Values are the mean of three independent experiments, with error bars representing the SD. (E) Representative FACS dot plots
showing apoptosis in cells of the indicated genotypes at 7 d postinfection or mock infection. Staining intensity for PI (x-axis) is plotted versus
that for Annexin V-FITC (y-axis). The percentage of cells categorized as apoptotic (top left quadrant; Annexin V positive, PI negative) or
necrotic (top right quadrant; Annexin V positive, PI positive) is indicated.
Conditional inactivation of Hus1 results in impaired cell proliferation and increased apoptosis. (A) Analysis of cell proliferation.
Regulated Hus1 Deletion in Primary Cells
Vol. 18, March 20071047
Page 5
both 4 and 6 dpi a small population of cells that possessed an
S-phase DNA content but were BrdU negative (designated
S2) was consistently observed specifically in Ad-cre–infected
Hus1flox/?1cultures (5.38% at 4 dpi and 4.61% at 6 dpi). These
abnormal S-phase cells were rare in control Ad-cre–infected
Hus1flox/?cultures (1.40% at 4 dpi and 1.50% at 6 dpi) as
well as uninfected cultures. The distribution of cells in other
stages of the cell cycle was largely unaffected by Hus1 loss.
A slight accumulation of Ad-cre–infected Hus1flox/?1MEFs
in G2/M was noted, but similar results were observed for
Ad-cre–infected Hus1flox/?MEFs.
The contribution of apoptosis to the reduced doubling of
conditional Hus1 knockout cells was also investigated. Ap-
optosis was measured by flow cytometric analysis of cells
stained with Annexin V, a phospholipid binding protein that
can be used to identify cells in which phosphatidylserine has
translocated from the inner to the outer leaflet of the plasma
membrane, a marker of apoptosis (Vermes et al., 1995). The
cells were also stained with PI as a measure of membrane
integrity, to distinguish intact cells in the initial stages of apo-
ptosis from cells undergoing necrosis or other forms of cell
death. The percentage of cells in the early stages of apoptosis
(Annexin V?PI?) in conditional Hus1 knockout and control
cultures is shown in Figure 2D. Beyond 4 dpi, Ad-cre–infected
Hus1flox/?1cultures consistently contained a greater percent-
age of apoptotic cells than Ad-cre–infected Hus1flox/?and
mock-infected Hus1flox/?1cultures, although these differ-
ences were not statistically significant. For example, at 7 dpi
12.3 ? 2.5% of Ad-cre–infected Hus1flox/?1cells were apopto-
tic, versus 9.0 ? 2.1% of Ad-cre–infected Hus1flox/?cells or
9.5 ? 1.1% of mock-infected Hus1flox/?1cells. Representative
plots showing FACS analysis of apoptosis in conditional
Hus1 knockout cultures are presented in Figure 2E. To-
gether, the results suggest that Hus1 inactivation causes
reduced cell doubling through subtle cell cycle alterations
and increased apoptosis.
Spontaneous Chromosome Abnormalities after
Conditional Inactivation of Hus1
Hus1 acts in a pathway that is essential for the maintenance
of genomic integrity. To quantify the extent of genome dam-
age in conditional Hus1 knockout cells, we determined the
frequency of gross chromosomal abnormalities in meta-
phase spreads prepared from Ad-cre–infected Hus1flox/?and
Hus1flox/?1MEFs. Ad-cre–infected Hus1flox/?1cells displayed
a dramatic increase in chromosome abnormalities that was
first detectable at 5 dpi (Table 1). By this time point, 55% of
conditional Hus1 knockout cells had at least one abnormal-
ity, and 45% had greater than two abnormalities, including
several cells with such extensive genome damage that it
could not be quantified. By contrast, only 11.8% of control
Ad-cre–infected Hus1flox/?MEFs contained chromosomal
abnormalities at 5 dpi, and none had greater than two ab-
normalities. Chromosome breaks and gaps affecting a single
chromatid were the most common abnormalities in condi-
tional Hus1 knockout cells (Figure 3A and Table 1). Chro-
matid interchanges were also observed at a low frequency.
To further characterize the spontaneous genome damage
that occurs after regulated Hus1 inactivation, we performed
immunofluorescence assays to detect ?-H2AX, the phos-
phorylated form of histone H2AX that accumulates at DSBs.
Although H2AX phosphorylation requires checkpoint sig-
naling, previous studies indicated that Hus1 is dispensable
for ?-H2AX accumulation after replication stress (Ward and
Chen, 2001). As shown in Figure 3B, there was a slight
increase in staining for ?-H2AX in Hus1flox/?1cells at 4 d after
Ad-cre infection, and by 7 dpi 16.9 ? 3.9% of Ad-cre–
infected Hus1flox/?1cells contained ?10 ?-H2AX–positive
foci, compared with only 3.1 ? 3.4% of Ad-cre–infected
Hus1flox/?MEFs. Mock-infected Hus1flox/?and Hus1flox/?1
cells displayed a low background level of ?-H2AX staining
similar to that observed for Ad-Cre infected Hus1flox/?cells
(data not shown). Thus, Hus1 loss specifically results in
increased H2AX phosphorylation and the appearance of
chromosomal abnormalities. We next investigated whether
these DNA lesions arose in a particular stage of the cell
cycle. This was accomplished by bivariate FACS analysis
of mock-infectedand Ad-cre–infected
Hus1flox/?1cells after staining with anti-?-H2AX antibody
and PI. The results confirmed increased H2AX phosphory-
lation in Ad-cre–infected Hus1flox/?1cells and further indi-
cated that the ?-H2AX accumulation after Hus1 loss was
largely restricted to cells with S-phase DNA content (Figure
3C). Importantly, this finding was not due to an inability of
this method to detect ?-H2AX in other stages of the cell
cycle, because positive staining was observed in G1, S, and
G2/M populations after treatment of cells with exogenous
genotoxins (data not shown; Marti et al., 2006). In sum, these
results indicate that conditional Hus1 inactivation causes
Table 1. Conditional inactivation of Hus1 results in increased chromosomal abnormalitiesa
Genotype dpib
Total cellsChromosome ChromatidTotal Avg.TotalAvg.
aMetaphase chromosomes were prepared from cells of the indicated genotypes, and the occurrence of chromosomal abnormalities was
bDays post-Ad-cre infection.
cExtensive damage refers to metaphases that contained too many chromosome abnormalities to be counted.
M. Zhu and R. S. Weiss
Molecular Biology of the Cell1048


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Ginger Borgeson sleeps in her vehicle at a road block in Buckhorn... News PhotoArts Culture and Entertainment,Colorado,Crystal Mountain,Fort Collins,Horizontal,Mode of Transport,Road,Sleeping,USAPhotographer AAron OntiverozCollection: Denver Post Ginger Borgeson sleeps in her vehicle at a road block in Buckhorn Canyon during the Crystal Mountain fire on Sunday, April 3, 2011. Borgeson said she was evacuated at around 12:30 a.m. when the fire grew from some 20 acres to more than 2,000 during the night. 'Better safe than sorry,' she said. 'I'd rather have 'em being concerned about us.' AAron Ontiveroz, The Denver Post (Photo By AAron Ontiveroz/The Denver Post via Getty Images)


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PowerColor Radeon R9 270X PCS+ 2GB Video Card Review - Pricing, Availability and Final Thoughts
Pricing, Availability and Final Thoughts
Coming in at $229.99 at the time of writing, the PowerColor R9 270X PCS+ 2GB sits in the middle of the pack when it comes to overall pricing. We've got a few cards that are more expensive, including the Devil 9 version of the card from PowerColor, which sits at $239.99. As always, when you start moving above the reference pricing, it's normally all about the cooler.
The PCS+ cooler that PowerColor uses seems to be strong. It puts out some great temperature numbers, albeit at a noise level that sits in the upper half of the graph. With that said, it's worth noting that while we are grabbing our noise levels, if we see the dB meter read something along the lines of 61 at first, and then finish up at 64.4 dB like it did today, the actual audible difference from 30cms away is not really noticeable. Of course overall though, we have to look at the numbers, and in the end this card does sit in the top half of the graph.
As for the bundle, there's almost nothing to it. With that said though, it is only slightly more sparse than what we normally see. The only thing this package lacks that we see most others include, are one or two Molex to PCIe power convertors. Considering how long PCIe power enabled power supplies have been around for though, we're not sure why companies still continue to include them in the package. The most important part of any package these days is the product itself, and we think PowerColor has done a good job in that regard.
When it comes to the performance side of things, PowerColor hasn't gone crazy with the clocks, but unlike a lot of other companies, they have chosen to increase the speed of the memory, which is nice. We can see that performance is slightly up over the reference card, which is as you'd expect due to the slight increase of the overclock.
When it comes to the Catalyst 13.12 Driver set today, there's clearly nothing in our benchmark line up that causes the driver to jump out in any huge way. Of course, with so many variations of the original 13.11 driver, it doesn't come as a huge surprise that there's not much change. During the course of that driver, we did see some decent changes, which saw big increases come to games like DiRT Showdown, and F1 2012.
If you're looking for a video card that comes in at that low $200 price bracket, then the R9 270X 2GB is a fantastic option. As with so many of the other R series cards, AMD has done a great job of pricing this card against its competition. Getting into specifics on what one to buy though, can be a little harder. PowerColor has put together a strong card here. It has an attractive cooler and while the overclock is not the largest, isn't bad at all; especially with the slight memory increase. And finally, the pricing is right where it needs to be.
PRICING: You can find products similar to this one for sale below.
United States: Find other tech and computer products like this over at Amazon's website.
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- Page 1 [Introduction and Package]
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As spring approaches, more law enforcement agencies are conducting DUI roadblocks or checkpoints. Many question the legality of the stops as they infringe on our rights to be free from unreasonable seizures. However, it isn't that easy. Oklahoma courts as well as SCOTUS have approved of the limited use of Oklahoma DUI checkpoints. This article explains Oklahoma DUI Checkpoints in more detail.
John Hunsucker
Oklahoma DUI Lawyer
Friday, April 10, 2015
Tuesday, March 31, 2015
How Oklahoma Cops Detect and Detain Suspected Drunk Drivers: Part Two
In our first post of this DUI Detection series, which covers the "DWI Detection and Standardized Field Sobriety Testing" guidelines for police officers, we covered the first phase ("vehicle in motion") of the three-phase process that police use when making the determination of whether to stop a driver on suspicion of driving while intoxicated. This post continues addresses the second phase, "personal contact."
We should note at the outset that the three-phase sequence -- vehicle in motion, personal contact and pre-arrest screening -- do not need to be followed in order. It is possible for a police officer to skip the first phase and go directly to the second; a common way this can happen is at a DUI checkpoint, or when the officer is responding to the scene of a car accident.
The purpose of the personal contact phase is for the officer to observe the driver while still in the car and to assess whether signs exist that suggest that the driver may be impaired. Depending on how this face-to-face interaction progresses, the officer will eventually decide whether the driver is sober, whether to go to the third phase and ask the driver to exit the vehicle so that field sobriety tests can be administered, or to arrest the driver based on probable cause of DUI.
When the officer begins the personal contact with the driver, he or she will be relying on sensory cues to assess the driver's state of sobriety. These cues include:
● Visual: The officer will be looking for intoxication-related symptoms such as bloodshot eyes, alcohol containers in the car, drugs or drug paraphernalia, unusual actions, soiled clothing, or shaky hands and fingers.
● Audible: The officer will be listening for the driver's admission that he or she has been drinking, slurred speech, inconsistent answers to questions, abusive language or unusual statements.
● Smell: The officer will note whether the odor of alcohol or marijuana is present, or if the driver has been using something like candies or breath sprays as an attempt to cover up such odors, as well as noticing whether other unusual odors are present in the vehicle.
Police officers are trained in interview techniques that impaired drivers will have difficulty with, most notably questions that require the driver to satisfactorily demonstrate divided attention tasks. For example, the officer may ask for the driver to do or produce two things at the same time, or may interrupt the driver's answer with another question (suddenly asking for the time or the date, or what the driver's middle name is), or using other techniques such as having the driver recite the alphabet or do a verbal countdown.
During this questioning, the officer will be looking for tip-off behaviors. For example, when asked for a driver's license and registration, does the driver forget to provide both, or provide a document other than one requested? Does the driver seem to be fumbling for the documents? This is a divided attention test whereby the officer is intentionally asking you to do several things at one time to see if you are able to comply.
If the questioning leads the officer to suspect intoxication, then he or she will usually instruct the driver to exit the vehicle. Even then the officer will be observing for symptoms, such as the driver's inability to follow instructions, inability to open the door, leaning against the vehicle or having to use the hands to steady his or herself.
In our next post in this series, we will go over what the officer may do in the third phase, the pre-arrest screening once the driver has exited the vehicle.
The Oklahoma DUI Lawyers as well as most of our support staff at the Hunsucker Legal Group are trained and certified in NHTSA DWI Detection and Field Sobriety Testing. Our goal is that we will be better trained and have more knowledge that the prosecutor and the police officer when we step into a courtroom to protect our clients. For a free consultation, please call 405-231-5600 or visit www.okdui.com
John Hunsucker
What does it really mean to be a convicted felon?
In Oklahoma, the trend has been to make more and more petty crimes into felony charges. A second time marijuana conviction in Oklahoma is a felony. Previously, a previous conviction for DUI could result in felony DUI arrest. Now, a previous deferred that has been expunged can be used to make a new DUI charge received 19 years after the first one a felony. The list goes on and on. the result is we are making our state a state of felons. Here are the consequences of being a convicted felon:
John Hunsucker
Oklahoma DUI Lawyer
John Hunsucker
Oklahoma DUI Lawyer
Tuesday, March 24, 2015
New articles dealing with breath testing and overcriminalization.
Just posted several new blog articles dealing with a couple different issues.
Recently, the Unites States Supreme Court (SCOTUS) recognized the term overciminalization in their opinion Yates v. United States. Basically, SCOTUS looked at the issue of prosecutors over reaching and attempting to stretch legal definitions to increase the potential punishment. Here is link to the blog article: http://oklahoma-criminal-defense-lawyer.com/u-s-supreme-court-ready-recognize-idea-overcriminalization/
The other article was posted on a national DUI blog that I also am a contributor. It was posted there as it is applicable to breath tests all over the nation. The article explains the issues with breathalyzers and Intoxilyzers. The article is posted here: http://duinewsblog.org/2015/03/24/factors-affecting-oklahoma-intoxilyzer-breathalyzer-tests/
Recently, the Unites States Supreme Court (SCOTUS) recognized the term overciminalization in their opinion Yates v. United States. Basically, SCOTUS looked at the issue of prosecutors over reaching and attempting to stretch legal definitions to increase the potential punishment. Here is link to the blog article: http://oklahoma-criminal-defense-lawyer.com/u-s-supreme-court-ready-recognize-idea-overcriminalization/
The other article was posted on a national DUI blog that I also am a contributor. It was posted there as it is applicable to breath tests all over the nation. The article explains the issues with breathalyzers and Intoxilyzers. The article is posted here: http://duinewsblog.org/2015/03/24/factors-affecting-oklahoma-intoxilyzer-breathalyzer-tests/
Tuesday, March 17, 2015
Oklahoma Expired Blood Test Kits: Is It Really Acceptable To Keep Using Them?
One of the tools that Oklahoma law enforcement use to gather evidence of driving under the influence (DUI) is a blood collection sample kit. This kit contains vacuum-sealed tubes, which are designed to draw in a pre-determined amount of blood which then mixes with an inorganic salt compound consisting of sodium fluoride and potassium oxalate.
Oklahoma blood test kits carry expiration dates on the vacuum-sealed tubes. There may be some confusion as to the effect these expiration dates may have if a police officer uses a tube that is beyond its expiration.
The first question is whether the expiration relates to the salt compound contained in the tube. The answer here is, "no." The compound itself is stable. This leads to the second, and more significant inquiry: if the expiration date on the blood collection vacuum tube does not refer to the salt compound, then what does it refer to?
The expiration refers to the tube manufacturer's warranty that the vacuum seal of the tube will maintain its integrity to a degree sufficient to draw a pre-determined amount of blood. It is the vacuum that draws the blood into the tube; but unlike the salt compound, the tube seal eventually does degrade and as it does, the amount of blood that it will draw into the tube becomes increasingly subject to uncontrolled variation.
So what does this mean to you, if you are subject to a blood draw using an expired vacuum tube?
The most significant impact of a loss of vacuum is that the amount of blood drawn into the tube may affect the ratio between the blood and the salt compound, which in turn may compromise the value of the test result when it comes to establishing whether a driver was under the influence when the blood sample was drawn. This potential volume-based skewing of the test sample is referred to as, "salting out".
A second possible problem with a loss of vacuum integrity in the collection tube is that a compromised seal means that more than the blood sample may possibly be able to enter it. Certain commonly-found yeast bacteria, for example, may be able to enter via the degraded seal. That bacteria consumes the blood sugar, or glucose, and as a waste product produces… alcohol. That is correct: a contaminated sample due to a faulty vacuum seal can result in the production of alcohol inside the tube.
What is more, there is no way for the devices used to analyze the blood sample for alcohol content to distinguish between the alcohol that was in your blood when the sample was drawn and any bacterially-produced alcohol that was created after the draw. This can lead to a challenge of the evidentiary value of the sample itself.
Are expired blood test kits being used in Oklahoma? The answer appears to be, "Yes." Documentation available from the state's Board of Tests for Alcohol and Blood Influence indicates that not only are some expired test kits still in the field, but that the State Director of the Board of Tests has given the green light to keep using them.
Not every blood draw sample in Oklahoma will be subject to vacuum-seal compromise. And the state is encouraging law enforcement in the state to turn in expired test kits. But if you have been accused of DUI based in part on the result of a blood sample, the fact that some of these expired kits are apparently still in use is an avenue that your attorney will definitely need to investigate as part of your defense.
Thursday, March 12, 2015
How Oklahoma Police Officers Detect and Detain Suspected Drunk Drivers: Part One
When it comes to making DUI stops, it is well-established in Oklahoma and other states that police officers cannot arbitrarily stop vehicles and then "work backwards" to try to find something illegal. The stop itself must have some basis in reasonable suspicion that the driver is driving while under the influence. But how do police make such a determination? Is there any uniform standard that they adhere to, and if so what is that standard?
Police in many jurisdictions do in fact have guidelines to help them decide whether to pull over a vehicle the driver of which they suspect is intoxicated. One such set of guidelines is issued by the US Department of Transportation's National Highway Traffic Safety Administration (NHTSA), and is entitled "DWI Detection and Standardized Field Sobriety Testing." It is important to realize that this manual is more than just how to perform the roadside field sobriety tests but also trains an officer when he should stop a vehicle suspected of DUI or DWI.
The NHTSA standards visualize a police DUI stop as a three-phase process that begins with observing the vehicle in motion, then progresses through making personal contact with the driver and finally to pre-arrest screening. This post will address the first of these three phases, "vehicle in motion."
This first phase is intended to answer the question, "Should I stop the vehicle?" Depending on the circumstances, the officer will answer the question in one of three ways:
● Stop the vehicle right away
● Wait and look for additional evidence
● Do not stop the vehicle
Driving a car involves a process known as "divided attention." A sober driver must be able to handle multiple tasks simultaneously, or near-simultaneously, such as steering, signaling, controlling the accelerator and brakes, and observing other cars and traffic signals.
Impaired drivers have trouble dividing their attention; they tend to concentrate on only a few critical tasks at any given time, letting the other tasks slip. These drivers frequently demonstrate symptoms that police are trained to look for, such as slowed reactions, poor coordination, impaired vision or impaired judgment.
There are, in fact, more than 100 driving cues that may indicate DUI behaviors, which fall into broader categories:
● Problems maintaining proper lane position
● Speed and braking problems
● Vigilance problems
● Judgment problems
Thus, police officers in phase one will be observing the vehicle to see if the driver exhibits behaviors like committing a moving traffic violation, swerving or weaving in a lane or across lanes, turning with an unusually wide radius, or driving at an unusually slow speed or with varying speeds.
The NHTSA standards even suggest the possible level of alcohol intoxication that these behaviors may indicate:
● Slowed reactions: blood alcohol concentration (BAC) of 0.03
● Impaired judgment: BAC of 0.05
● Impaired vision: BAC of 0.08
● Poor coordination: BAC of 0.10
Other things that police will look for include expired registration tags or movement in the car indicating that the driver is drinking something or otherwise taking drugs.
The officer's decision whether to move to the second phase -- stopping the vehicle and interacting with the driver -- can depend on the severity of the impairment symptoms that he or she sees. Dangerous behaviors, such as nearly striking other vehicles or objects or driving with one's headlights off at night will likely result in an immediate stop; less severe behaviors, like driving slowly or drifting within a lane may lead the officer to wait and see if the driver displays other symptoms.
In a following post, we will consider phase two of the three-phase process: the vehicle stop and the officer's communication with the driver.
Every one of our Oklahoma DUI Lawyers as well as most of our support staff at the Hunsucker Legal Group are trained and certified in NHTSA DWI Detection and Field Sobriety Testing. It is our goal that we will be better trained and have more knowledge that the prosecutor and the police officer when we step into a courtroom to protect our clients. For a frre consultation, please call 405-231-5600.
Monday, March 2, 2015
Field Sobriety Tests: How Reliable Are They?
Aside from Intoxilyzer or blood tests, one of the commonly-used means of Oklahoma police to assess whether an Oklahoma driver is under the influence of drugs or alcohol are field sobriety tests. Officers typically have a suspected drunk driver perform a battery of sobriety tests; failing one or more of them can not only lead to an arrest for DUI, it can possibly also be used by the prosecution as evidence at trial.
Field sobriety tests are not, however, foolproof as indicators of intoxication. To begin with, they are limited in their evidentiary value. None of these tests can be used as a correlation to a specific blood alcohol content level, because they may only indicate intoxication (and not necessarily alcohol intoxication). Moreover, even a test that may seem to suggest intoxication might not hold up in court if reasonable doubt can be cast on the way that police officer conducted it.
Precision is key. Just as an Intoxilyzer must be properly calibrated for its results to hold up in court, field sobriety tests must be administered correctly to be valid. Even seemingly minor mistakes by the officer during the testing can open the door to challenging the test results, and a successful challenge can make the difference between a conviction for DUI or an acquittal.
The National Highway Transportation Safety Administration (NHTSA) establishes the requirements to properly administer field sobriety tests. These guidelines are quite precise; the inadvertent failure of a police officer to follow them properly is a definite possibility.
As one example, consider the "horizontal gaze nystagmus" test. This test involves instructing the test subject and then having the test subject follow the track of a stimulus object, such as a flashlight, with his or her eyes while holding still. In theory the test sounds simple to administer, but there are several ways that the police officer can commit errors that invalidate it.
The test itself consists of four phases and 14 passes of the stimulus object. The NHTSA requirements are specific on how the test subject is to be positioned, how far from his or her nose the stimulus object should be held, how quickly it should be moved, the maximum angle of deviation that should be used, and the minimum time that should be spent conducting the test; if it takes less than 86 seconds then it is suspect because the officer has performed it too quickly, which in turn shows that one or more of the steps was administered correctly.
For anyone accused of DUI in Oklahoma based at least in part on the results of field sobriety tests, what is important to know is that not only is it imperative that the police officer who administered the test followed the correct procedure with exactitude, it is also essential that the Oklahoma DUI defense attorney also be intimately familiar with the details of the test protocols. Only an experienced DUI attorney who has carefully studied the NHTSA requirements for how a valid test should be given will know what to look for when it comes to identifying ways in which the officer may have made a mistake. The DUI Attorneys at the Hunsucker Legal Group have received the same field sobriety testing training as the police officer on the street and are versed in defending Oklahoma drivers accused of DUI or APC. Most often, they know the Field Sobriety Manual better than the arresting officer.
John Hunsucker
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... Have I mentioned how **** the coffee is in America? It doesn't even taste like coffee & is usually served straight from a press button machine. Makes you realise how spoilt we are in Australia & New Zealand with good coffee. I've been mainly drinking tea. Walked through the town & ordered two cappuccinos that were half the price & was stoked that it tasted great for once! The ...
South Beach Baby!!!
... our bags and then tried to figure out how we'd get from Ft. Lauderdale to Miami beach. We were initially going to give mass transit a try and use a combo of trains & busses, but decided on a shared ride shuttle can at the last minute. Great choice! They upgraded us to a town car instead. Nice! The ride was quick and we passed our cruise ship on the way to the hotel. We were very early, so we dropped our bags off at The Clay Hotel and went off to ...
A slow start to something possible extraordinary
... for me. I too am for her as she's making progress with her flat purchase, I'll be soooooooo pleased to see her with her own place, she deserves it. Perhaps now we really are just good friends which is brilliant.
After all of this I calmed myself down with a walk to the beach, it was amazing watching the colours of the cloud lit off the sun. I then walked along the beach and saw a group of people taking pictures of them as a group ...
Boredom is...
... always such a treat :) This morning we got up and Ellen trained me to be a perfect stew. It was a day's work, so happy to have Ellen here! I took notes and watched and learned. The owner of the boat phoned me and we also had a good chat, he will phone one of my au pair references tomorrow and will then hopefully get an answer. I went home at about 5 and took a nice, long relaxing bath. Nicole and Kate have left :( All us housemates had a braai. Holding thumbs for ...