text:
Supported by a grant from the National Natural Science Foundation of China (No 30772460).
A LDR-PCR Approach for Multiplex Polymorphisms Genotyping of Severely Degraded DNA with Fragment Sizes <100 bp*
Article first published online: 5 OCT 2009
DOI: 10.1111/j.1556-4029.2009.01166.x
© 2009 American Academy of Forensic Sciences
Journal of Forensic Sciences
Volume 54, Issue 6, pages 1304–1309, November 2009
Additional Information
How to Cite
Zhang, Z., Wang, B.-J., Guan, H.-Y., Pang, H. and Xuan, J.-F. (2009), A LDR-PCR Approach for Multiplex Polymorphisms Genotyping of Severely Degraded DNA with Fragment Sizes <100 bp. Journal of Forensic Sciences, 54: 1304–1309. doi: 10.1111/j.1556-4029.2009.01166.x
- †
Publication History
- Issue published online: 23 OCT 2009
- Article first published online: 5 OCT 2009
- Received 18 June 2008; and in revised form 13 Oct. 2008; accepted 2 Dec. 2008.
- Abstract
- Article
- References
- Cited By
- forensic science;
- DNA typing;
- DNA degradation;
- rs17750303;
- rs2307647;
- rs2307557;
- rs17250992;
- ligase detection reaction;
- polymerase chain reaction
Abstract: Reducing amplicon sizes has become a major strategy for analyzing degraded DNA typical of forensic samples. However, amplicon sizes in current mini-short tandem repeat-polymerase chain reaction (PCR) and mini-sequencing assays are still not suitable for analysis of severely degraded DNA. In this study, we present a multiplex typing method that couples ligase detection reaction with PCR that can be used to identify single nucleotide polymorphisms and small-scale insertion/deletions in a sample of severely fragmented DNA. This method adopts thermostable ligation for allele discrimination and subsequent PCR for signal enhancement. In this study, four polymorphic loci were used to assess the ability of this technique to discriminate alleles in an artificially degraded sample of DNA with fragment sizes <100 bp. Our results showed clear allelic discrimination of single or multiple loci, suggesting that this method might aid in the analysis of extremely degraded samples in which allelic drop out of larger fragments is observed.
The breakdown of genomic DNA into small fragments by chemical or physical factors is always a special challenge in the genetic identification of human remains or crime stains (1). One strategy that addresses some of the problems encountered in analyzing degraded DNA is the reduction of polymerase chain reaction (PCR) amplicon size (2). Mini-short tandem repeats (STRs) and single nucleotide polymorphisms (SNPs) are two types of genetic markers that can be analyzed in an amplicon smaller than 150 bp (3). A number of validated mini-STR and SNP assays have been used in casework in which current STR kits failed to achieve complete DNA profiles (4–9). However, these two techniques require an average DNA fragment length of 100–200 bp and may not be applicable to casework with extremely degraded DNA consisting of fragments <100 bp in length.
Ligase detection reaction (LDR) has been used to detect mutations in disease genes in clinical samples (10–13) and provides an elegant technique for polymorphism typing that could be applied to severely degraded DNA. Here we present a multiplex polymorphism typing method that couples LDR with PCR (LDR-PCR). In the LDR portion of the technique, probes that are designed to be complementary to the polymorphism and to the region directly adjacent are ligated when brought into juxtaposition through hybridization with the DNA fragment. In the subsequent PCR step, the ligation products are amplified using primers that are complementary to the generic priming sites built into the hybridization probes (Fig. 1). In this study, the LDR probes were designed such that only 20–40 bp of intact DNA was required for their hybridization. These techniques were applied to samples of artificially degraded DNA consisting of fragments <100 bp to determine whether each allele of four different polymorphisms could be discerned.
Materials and Methods
Selection of Polymorphic Markers
The polymorphic markers used in this study were selected from the dbSNP database (http://www.ncbi.nlm.nih.gov/SNP/) and included two single-base nucleotide substitutions and two three-base deletions/insertions (Table 1). These markers were chosen because they met certain empirical criteria including randomness of nucleotides in the target region, low secondary structure of the target sequence, and uniqueness of the target sequence within the whole genome. The exact chromosomal locations were determined using BLAST (http://www.ncbi.nlm.nih.gov/blast/Blast.cgi).
|Chromosome Position
Oligonuleotide Primers and Probes
All oligonucleotide primers and probes were synthesized by Takara Biotechnology Co., Ltd. (Dalian, China) and Shanghai Sangon Biological Engineering Technology and Services Co., Ltd. (Shanghai, China). Primer and probe sequences were given in Tables 2 and 3. The DNA sequences for design work were downloaded from the National Center for Biotechnology Information (NCBI) website (http://www.ncbi.nlm.nih.gov/).
|Sequence (5′3′)
|LDR Probe*
|Total Sequence (5′3′)†
|Length of Target-Specific Region (nt)
|Probe Length (nt)
|LDR-PCR Amplicon Size (I+II/III, bp)
|Generic primers for PCR of ligation products
|Primer 1
|Primer 2
Preparation of Genomic DNA Samples and Genotyping of the Polymorphic Loci
Twelve anonymous liquid blood samples were provided by the Department of Forensic Serology, China Medical University (Shenyang, China). Genomic DNA was extracted by the standard phenol–chloroform procedure and was quantitated based on UV spectrophotometry (A260). To avoid any possible contamination, all procedures for DNA isolation were carried out in separate rooms using dedicated equipment.
Prior to degradation, all 12 genomic DNA samples were genotyped at each of the four polymorphisms studied. DNA was amplified with each set of sequencing primers (Table 2). The PCR products were purified and sequenced by Shanghai GeneCore BioTechnologies Co., Ltd. (Shanghai, China). The genotypes of each of the 12 samples for the four polymorphic loci were identified according to the sequencing results.
Preparation of the Reference Sample of Artificially Degraded DNA
We subjected the genomic DNA samples to DNase I digestion, then assessed the severity of degradation by PCR using primers 100 bpF/R (size of PCR products: 102 and/or 105 bp). Each 90 μL digestion reaction consisted of 9 μL of DNase I (1 U/μL) (Fermentas, Vilnius, Lithuania), 9 μL of 10 × DNase I reaction buffer without MnCl2 (100 mM Tris–HCl [pH 7.5 at 25°C], 1 mM CaCl2), 9 μL of 100 mM MnCl2, and 10 μg genomic DNA, incubated at 37°C. Nine μL aliquots were removed at 0, 10, 30, 60, 90, 120, 150, 180, 210 min time points, and DNase I activity was quenched by adding 1 μL of 25 mM EDTA with heating at 65°C for 15 min.
The extent of DNA fragmentation was assessed by PCR amplification using primers 100 bpF (5′-GGGAGCCCACTGACTGAAC-3′) and 100 bpR (5′-GTCTCCAGAAAAGGAGGACATC-3′) performed in a Tgradient thermocycler (Whatman Biometra, Goettingen, Germany). Each 20 μL PCR reaction contained 0.1 μg of DNase I digested DNA, 0.2 μM 100 bpF, 0.2 μM 100 bpR, 200 μM dNTPs, 1 U of rTaq DNA polymerase (Takara Biotechnology Co., Ltd.), and 1 × PCR buffer. Reaction parameters consisted of an initial denaturation step at 94°C for 2 min, followed by 34 cycles of 94°C for 30 s, 60°C for 20 s, and 72°C for 20 s, with a final extension at 72°C for 3 min. PCR products were separated on a 3% polyacrylamide gel and visualized by ethidium bromide staining. Samples too degraded to be amplified were used as the reference sample for the subsequent LDR experiments.
Ligase Detection Reaction Coupled with PCR
Each 20 μL ligation reaction contained 4 U of Ampligase® Thermostable DNA Ligase (Epicentre Technologies), 1 × Ampligase® Reaction Buffer, 50 ng of DNase I digested DNA, and 1 or 2 pmol of the LDR probes (see legends for Figs. 3–5). Following an initial denaturation at 94°C for 3 min, the ligation parameters consisted of 20 cycles of denaturation at 94°C for 30 s, and ligation at 37, 42, or 45°C for 8 min (see legends for Figs. 3–5). Different probe concentrations and ligation temperatures were used depending on whether a polymorphism was being determined individually (Fig. 3) or in a multiplex reaction (Figs. 4 and 5).
PCR reactions were performed using the ligation product of the LDR as amplification template. Each 30 μL reaction contained 3 μL of the LDR product, 200 μM each dNTP, 0.2 μM each generic primer (Table 3), 1 U rTaq polymerase (Takara Biotechnology Co., Ltd.), and 1 × PCR buffer. The PCR parameters consisted of an initial denaturation at 94°C for 3 min, followed by six cycles of 94°C for 30 s, 54°C for 20 s, and 72°C for 15 s, followed by 28 cycles of 94°C for 30 sc, 50°C for 20 s, and 72°C for 15 s, and a final extension at 72°C for 5 min. The products were separated on a 10% polyacrylamide gel and visualized by ethidium bromide staining.
Results and Discussion
Preparation of the Reference Sample of Degraded DNA with Fragment Sizes <100 bp
In crime case investigations, DNA samples are often qualitatively inadequate since the degradation of human DNA varies with light, humidity, elevated temperatures as well as bacterial and fungal contaminations followed by the growth of these microorganisms (1). To systematically assess the performance of different typing techniques, it would be desirable to have a standardized reference sample of degraded DNA in sufficient amount and specific length (14). Artificially degraded DNA can be obtained by sonication, enzyme digestion, or physical methods such as elevated temperature or humidity (3,15,16). DNase I digestion for controlled production of degraded DNA has been described in several studies (15–18).
In the presence of Mn2+, DNase I enzyme cleaves double-stranded DNA and completely digested products are tetranucleotides (19). Increasing incubation times with DNase I resulted in increasing severity of degradation. Analysis by agarose gel electrophoresis (data not shown) showed that the sample incubated for 0 min consisted of high molecular weight DNA, and samples incubated for 10, 30, and 90 min consisted of fragments that were approximately 600, 200, and 100 bp, respectively. However, since the sizes of the degraded DNA fragments could not be reliably assessed by agarose gel, assessment by PCR amplification was used as described previously (16,20). We designed a pair of primers (100 bpF/R) and attempted to amplify a 102/105 bp amplicon using the degraded DNA as template. In order to ensure that there were no detectable large DNA fragments remaining, we increased the quantity of DNA template (0.1 μg of DNase I digested DNA) and the cycle numbers of PCR (34 cycles). With increasing digestion time, we observed decreasing amounts of PCR product, with no amplification product from the DNA samples digested for 150 min or more (Fig. 2). As an additional check, we attempted to amplify 8 ng genomic DNA digested for 150 min using the AmpFlSTR® Identifiler® PCR Amplification Kit (Applied Biosystems, Foster City, CA) according to the manufacturer's instructions. Analysis of this reaction gave no results (data not shown). Therefore, we were confident that the length, and not the quantity, of DNA fragments caused the PCR reactions to fail, and that the 150 min digestion sample could be used as the reference sample of degraded DNA with fragment sizes <100 bp. In our experiments, every DNA digestion sample was verified to contain fragments under 100 bp by failure of PCR using primers 100 bpF/R before use in subsequent LDR experiments.
Rationale for Using LDR-PCR and Selection of Polymorphic Markers
A forensic DNA sample frequently is too degraded to give a full profile of STRs. Genotyping methods that can be successfully performed on smaller fragments of genomic DNA are of great interest to forensic science. Reducing the amplicon sizes of target DNA fragments has become a major strategy of analyzing the degraded DNA, and mini-STR-PCR and mini-sequencing techniques have been used in casework (5,6,8). However, due to the length limitation of PCR amplicons (21,22), these PCR-based assays would not provide reliable DNA profiles from a sample of severely degraded DNA such as our reference sample (DNA digested with DNase I for 150 min).
To characterize a bi-allelic polymorphism by LDR, three probes (one common and one specific to each allele) must be designed (Fig. 1). When allowed to hybridize to the target DNA, the probes will be brought into juxtaposition and ligated by DNA ligase. The probes that discriminate the two alleles (probes II and III) have, as their 3′ terminus, the nucleotide(s) corresponding to their target allele. This strategy capitalizes on the sensitivity of DNA ligase to all base mismatches on the 3′-side of the nick (23). Only probes that hybridize completely become ligated by the thermostable ligase used in this study. The two probes together only hybridize to approximately 20–40 bp of target sequence, making a degraded DNA sample an adequate template. After several cycles of hybridization and ligation, the ligation products would be sufficient for the subsequent PCR amplification.
Each LDR probe was designed to contain a generic primer-specific sequence (Table 3) so that every set of LDR-ligated product could be simultaneously amplified in the PCR step (Fig. 1). In order to avoid nonspecific priming, the sequences added to the LDR probes used for priming the PCR reactions were derived from E. coli plasmid replication origin and adjusted by ourselves. In addition, several nucleotides were added between the primer-specific and target-specific regions of the probes so that all PCR products of ligated LDR probes varied in length by 3 bp and could be discerned by polyacrylamide gel electrophoresis (Table 3 and Fig. 1). Together, these measures reduce the risk of amplification artifacts and provide for low cost, high throughput analysis of multiple polymorphic loci.
Our next challenge was to identify polymorphic markers that could be accurately distinguished by the LDR-PCR method in a degraded DNA sample. In the interest of sample conservation, it was also important that the design accommodate multiplexing of the reactions. Using the dbSNP database, we identified two SNPs (rs17750303 and rs17250992) and two three-base deletion/insertion polymorphisms (rs2307647 and rs2307557) that met our criteria and were selected for investigation (Table 1).
Genotyping of the Reference Sample of Degraded DNA by LDR-PCR
After genomic DNA was digested with DNase I for 150 min, DNA fragments were less than 100 bp in length (Fig. 2). Despite the severe degradation of the DNA samples, each of the four polymorphic loci were successfully discriminated using our LDR-PCR method (Fig. 3). LDR capitalizes on the ability of DNA ligase to covalently join two adjacent oligonucleotides hybridized to target DNA in which there is perfect complementarity at the nick junction (13). Without pre-amplification of the target sequences, our LDR probes could directly recognize and hybridize their targets within the degraded reference sample. Therefore, only a very short target sequence (20–40 bp surrounding the polymorphism) within the DNA sample was required for the ligation reaction. LDR-PCR is therefore a genotyping technique suitable for use in extremely degraded DNA samples.
Studies of the recognition process of DNA ligases show that when binding to the DNA substrate, their footprints extend approximately 5–9 nucleotides on the 3′-OH side of the nick, and 9–12 nucleotides on the 5′-phosphate side of the nick (24,25). Thus, DNA ligases require a certain length of double stranded DNA template in order to stably bind to the nick and catalyze the repair reaction. The design of LDR probes rs2307557 and rs17250992 included 17–20 bp complementary to the target DNA (Table 3), which would provide more than sufficient double stranded DNA for ligase recognition and binding. However, the primary goal of this LDR technique was to detect polymorphisms in severely degraded DNA samples containing very small fragments. Therefore, for the other two polymorphisms, the LDR probes were designed with shorter target-specific sequences (rs17750303 and rs2307647, Table 3), containing only 10–13 bases complementary to the target region. The results showed that Ampligase® allowed for successful ligation reaction and polymorphism discrimination when the target-specific sequence of LDR probe was as short as 10 nt (Fig. 3A, B). This characteristic of Ampligase® is valuable in this type of analysis.
While most DNA ligases have very stringent requirements for perfect complementarity on the 3′-side of the nick, it has been reported that T–G and G–T mismatches at the 3′-end of the nick can generate a quantifiable level of ligation product (23). A ligation product formed despite a sequence mismatch would result in unreliable typing because erroneous ligation products would also be amplified in the subsequent PCR step. In this study, the rs17250992 polymorphism is a C/T SNP and therefore there was the possibility that the probe used to discriminate the "C" allele would erroneously bind to the DNA template having the "T" allele, producing a G–T mismatch at the 3′-end of the nick. Despite this situation, this polymorphism was accurately discriminated (Fig. 3D). For each polymorphism studied, two allele-discriminating probes competed to anneal to DNA template adjacent to the common probe, yet we observed that only the allelic probe with perfect complementarity to the template became ligated to the common probe by Ampligase®. This high degree of allele discrimination might result from the use of short LDR probes (23), intermolecular recognition and fidelity of Ampligase®.
Multiplex LDR-PCR Reaction
Design of a multiplex LDR-PCR reaction requires similar annealing temperatures for all probes and an avoidance of interaction effect between them. When designing the probes for the LDR-PCR assays, Tm of each LDR probe was calculated using the online HyTher software (26). We then tested the specificity of each pair of probes (probes I+II or probes I+III) by performing the hybridization step at different temperatures (the Tm, and 5°C above and below the Tm). PCR amplicons of the ligation products from each hybridization were similar (data not shown), therefore we were confident that the specificity of the reaction would remain even in the presence of slight fluctuations in hybridization temperature. Additionally, Primer Premier 5.0 software was used to analyze any interaction between the LDR probes, which found no significant risk of mis-hybridization between probes and target loci.
We then mixed all four sets of probes and carried out the LDR-PCR procedure in a single tube. Based on the fact that the target-specific oligos of LDR probes were different in length and sequence, the ligation temperatures and concentrations of probes were modified experimentally. Using 2 pmol of each probe for rs17750303 and rs2307647, and 1 pmol of each probe for rs2307557 and rs17250992 at a hybridization temperature of 42°C, we correctly genotyped four samples of DNase I digested DNA in a multiplex LDR-PCR reaction (Fig. 4). All alleles of the four different loci were readily distinguished, confirming a lack of interference between probes.
Finally, we confirmed the genotypes of all 12 reference samples of digested DNA at the four polymorphic loci using our multiplex LDR-PCR method. The 12 samples had been genotyped by direct DNA sequencing of the polymorphic regions before digestion with DNase I. The digested samples were then randomly coded by a researcher who was blind to the original genotyping results. The coded samples were then genotyped using the LDR-PCR method. After analysis, the code was broken, revealing that the genotypes of each sample for each of the four polymorphic loci had been correctly identified (Fig. 5). Therefore, this LDR-PCR method can be used to perform multiplex genotyping of polymorphic loci in samples of severely degraded DNA with fragment sizes less than 100 bp.
Indeed, SNPs and small-scale multi-base deletions/insertions are not as powerful as STRs for identification purposes, but they can provide valuable information in cases of highly degraded samples that yield incomplete STR profiles or no profile at all (3,27,28). This pilot study confirmed the potential of LDR-PCR methods in genotyping polymorphic loci in severely degraded DNA samples. Future work should focus on selection of target polymorphic loci, design of LDR probes, and optimization of reaction conditions such that dozens of loci could be discriminated simultaneously in a single tube. Additional modifications to the protocol, such as using florescent dyes for allele detection rather than ethidium bromide, will only improve the assay, making it sensitive enough for forensic casework.
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This dude is one lucky fuck, he gets sucked by 3 girls all of which squirt at an alarming rate. Then he ass and pussy fucks them and makes them squirt a lot more....They do
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Super sexy Sapphire dresses up as a school girl complete with pig tails. She then sucks on her glass dildo which she slides between her big tits before she brings it down and jabs it into her young tight shaved pussy.
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Exceptional Asian girl Marica Hase gets Danny Ds cock in her gentle mouth and hairy pussy
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Busty baseball star Devon Michaels is going to punish Johnny Sins for his stupid mistake after game in the locker room. She makes him lick and fuck her pussy. He likes the punishment.
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This girl is amazing. From her huge ass to her pretty dick sucking lips. There is plenty dick sucking, pussy licking, ass pounding, ass licking and cum swallowing for anybody with taste could love!
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I just love a girl with short hair. Dont ask me why. Its just sexy. Well, apparently, this older gentleman agrees, if his enthusiastic pussy eating is any indication. Nice!
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This short haired girl with large natural boobs came for audition. Her name is Tracy Lee! Tracy is very attractive girl and think that great sex with this hottie will be unforgettable.
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Start getting a lot of pleasure with White Angel right now and you would fall in love with her! The blonde girl is becoming naked before starting to play with sweet pussy.
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College boys better shape up or ship out. These girls are here to tuenthings up . We got Diamond Kitty who loves to torture the boys into doing shit they thought they'd ever do. Then we got the gorgeous Britney Amber who is holding some grip is slapping that thing around making all the boys feel in adequate (lets just say they don't match up). And finally the nastiest and sluttiest Porn super star to come along AVA FUCKING DEVINE! She is going to fuck all the boys in college ten at a time. I don't think there is a enough college for these three girls.
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Gorgeous Lesbians Sharing Toys and Strapon in Amazing Girl-on-Girl Vid
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The sexy girl Anikka Albrite possesses the real treasure in the view of the delicious looking big bubble butt that she willingly demonstrates wrapped in tight shorts and fishnets
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Stupid slut Donna Bell lets some hung guy spray cum on her mouth
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Lexie is a club girl, or at least that's where we picked her up. After we flashed a little cash she was willing to do anything to keep us happy. This girl gobbled cock like a pro and got her tight little slit slammed hard over and over, just for a bit of the green. Imagine her fury when we covered her face with cum, had her sign the model release and then fucked off without paying her!
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Today we have girl who I think will have a long and prosperous career. Her name is Bonnie and she is a hottie! This girl is super flexible and loves to fuck! She has never had a cock that was really big tho, so we decided to let Jmac come and hang out wit
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Fer and Diego were walking to the studio when they have met a charming blonde girl. They have invited her to visit their work and drink some beer there. Thamara had absolutely nothing to do and was glad to waste some time with cute guys. She even thought it would be great to fuck him. They have came to the studio with pretty girl and beer and their friend has decided to shoot their drunk party on the camera. Thamara has refused Diego with his moves initially so guys have decided to add vodka to Thamara's beer to get her more relaxed. Thamara has drunk few glasses before she has wanted to fuck here and now. She even wanted both guys at once. Fer and Diego were surprised with the drunk passion of their new girlfriend. Thamara has sucked their cocks like crazy and opened her pussy and ass for big men's dicks. And she could not get enough of that..
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Irish beauty Shannon Reid and Czech sexpot Ashley Bulgari are back on Euro Girls on Girls, this time as sultry blondes and heating up the screen together in every man's girl-girl massage fantasy come true!
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Sometimes its hard for a girl to get fully satisfied by a man. In that case only the best friend can help her. Especially when the friend is a steaming hot lesbian!
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This is a one in a million girl. Beautiful girl with a HUGE ass. White chicks dont normally have an ass, and she has one of 3 girls. Her ass is gorgeous!
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If you're in to hot Girl-on-Girl squirt parties, then Flower Tucci is going to wet your whistle nicely in this scene with Angela Stone, Britney Stevens and Sasha Grey. Most of these fine young sluts show up wearing masks and flashy bikini's, but it doesnt
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Cameron is out new audition for this week. She came to us from Canada, to do some adult movies. Cameron already has experience in the industry as a web-cam girl and she's extremely flexable too, because Cameron is also a part-time Yoga instructor. Cameron is a very gorgeous blonde, with a very sexy body and she loves sex. I told her to demonstrate her skills on a dildo I had on the desk and Cameron really showed us why she was a perfect addition. She did things to that dildo that no other girl has ever done in this office. I wasn't done with her either, I still wanted to see how she fucked, So I called in the stud and thats when shit got really intense.
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Hi, it is the video from bang bus again. Today you could see how Gulliana Alexis is getting seduced to screw. This girl is just 18 year old and she wants to become a pornstar.
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The guys were on another quest looking for sexy amateur girls to pick up. They meet Alice and this 20 year old girl seems to know how to have fun and takes their invitation to take a ride.
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Stupid brunette slut Charity Bangs rides cock after 69 position
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Luana fernandez`s ravishing wazoo is worth tthis chab extra cAsh
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Hardcore lesbian action with Jennifer Annah Fox,Mercedes Lynn and Sofia Ressen. These babes tease each other on the beach and then go back to a flat where they quickly get stuck into some serious girl on girl action.
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So we got this video in... from somewhere.. I don't know where.. but who cares, it's got some HOT girls on it! These girls snuck into a dorm room at night while their rushes were sleeping and scared the shit out of them and then made them strip down to nothing and do jumping jacks and push ups. They stuck vibrators in their pussies and took them all to the showers to clean off. They made the rushes lick their pussies until they came! Damn the shower scene was pretty hot, the girls actually started to enjoy it and get off!
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Precious blonde girl Heather Starlet wearing a short skirt and sitting on the desk with wide open legs, and her naughty boyfriend is squatting all naked and hotly licking and sucking girls young cunt!
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Student girl Eufrat gets used by crazy bad teacher Megan Coxxx. Four-eyed brunette with small pierced tits spreads girls legs after removing her panties. She tongue fucks her pussy and then makes her eat her snatch!
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Three hot chicks hanging out and trying on lingerie, skimpy panties and lacy underwear. And when its Diamond Kitty, Mercedes Lynn and Victoria Voss then you know youre going to get some sexy, horny, girl girl action.
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This is why we love country girls! Just look at Athena Pleasures and youll realize that this babe is as sexy as country girls come with a massive set of boobs and a butt that was made for fucking!
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It's a new year and we want to start it off right!! We're going all out with more hot girls than you can imagine! We PACKED this strip club full of as many girls as the fire code would allow us to, and believe me, they all want a shot at the cock! Happy new year, and ENJOY!
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tight assed brunette teen girl Rita D was at picnic with her handsome boyfriend, at some time he started to gently fondle her hips and stroke her sexy tight boobies girl enjoyed the hot cunilingus and great fuck at the nature!
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What do you get when you put a hot blonde and a slutty brunette in the same room and give them a bunch of toys to play with This amazing girl on girl video with Jane and her girl!
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Have you ever fucked a Hawaiian girl No! Me neither. But after watching Mahina Zaltana take dick Im convinced that I want to fuck a Hawaiian girl. Mahina loves to suck this big black dick.
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A guy is driving along in his car. There is a teenage girl next to him and with one hand he strokes her legs. The the girl turns around and gives him a blow job. The both of them get out of the car and the both of them fuck on the ground.
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Handsome teacher Tommy Gunn seduces stupid coed Victoria Rae Black to spend cool time with him. Girlie starts sucking fat rod of man immediately before getting cunnilingus.
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Hot girl Amia Miley is with her boyfriend at a gorgeous restaurant eating their dinner. Of course the girl needs to pay her boyfriend back for this expensive food.
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Kissing HD Close-up tongue sucking with adorable young lesbian girls
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What a group of hardcore party animals we had at the this club. There was a couple shy girls, but they opened up after seeing how much fun all the other girls were having. I'm not sure who was enjoying it more the girls or the guys. Everybody was playing with everybody. The girls even set up a panty pyramid. Jmac was warming up a few girls, but Ahna is the one that got opened up wide. Her big titties were bouncing along with the big beats. Jmac made sure she got filled up with his cock and covered her is man sauce.
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We get a lot of beautiful girls here at Bangstudios. But sometimes, we get girls that are so incredibly hot that we just dont know what to do with ourselves. Take Alexa Rydell for instance. What a gorgeous woman. She came in and I knew I wanted to see her naked right away. Alexa is a sweet girl with a very naughty side to her and she wanted to get fucked right away. I was all for it. I Auditioned her and called in the stud to test her out. I think she has a bright future.
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Brunette horny wife cheats her stupid husband with her ex BF in the room. Hot video
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Asian Girl In Stockings Getting Her Asshole And Pussy Fingered By 3 Girls On The Massage Chair
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Cleopatra style girl gets her pussy licked by other girl in a bathtub
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Bree Daniels is tired from everything. Her work is boring, her husband is stupid. Only her pussy is with her and in gives lots a satisfaction.
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In this bangbus update we have the sizzling the gorgeous Rebecca and man does this girl deliver or hat she even picked some girls up from the crowd of people!I mean this girl can do it all!Enjoy!
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Theres nothing better than the introduction of a new girl on film.This girl we have today is really cute. What we do is teach girls about the biz and see if in the end of them getting fucked on camera wether its something theyd like to continue doing.
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This lucky stud scores two chicks when he is out clubbing one night. They go back to his hotel room and get busy on the bed; as his girls strip off, the guy seems to notice that there is something funny about one of them and that she has a peculiarly husky voice! When she pulls off her panties he is stunned to see a cock and balls staring him in the face. He finds himself curiously aroused by sharing a bed with a big dick shemale, especially when the other girl starts to go down on the tranny. There is a great amount of sexual variety in this clip, ranging from hardcore 69ing, to double blow jobs and hardcore anal penetration.
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The Shop Girls: Girl/Girl action in the Store Room with a Surprise Cock
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Somebody give this girl a detention because she has been very bad indeed! The big breasted Lola Lane is only too happy to teach this naughty school girl a thing or two. Rubbing that tight ass all over the her mistresses' lap really is asking for trouble, and Ms Lane gives those peachy butt cheeks a hard slap. She lifts up her short skirt so the other girls can play with her neat little twat. Things get really dirty as all three ladies take turns sucking on each other's pussies. Eager to please the little slut spreads her legs willingly so her tutors can finger and slap her cunt. This is a lesson in fucking she will never forget.


text:
How Feminist Is the Occupy Movement? – November 28, 2011 at 1:00pm
Support KPFA Today
Community Powered Radio for the Bay Area and Beyond
Founded in 1949 by Lewis Hill, a pacifist, poet, and journalist, KPFA was the first community supported radio station in the USA. Keep us going another 50 years!Secure Online Donations


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FaceOffSimon & Schuster 2014; US$ 26.99
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text:
Four machines demonstrated recycling applications at U.K. event.
Customers and distributors from over 21 countries attended the 2013 Powerscreen Open Day held on Oct. 3-4 at Arden Brickworks near Birmingham, U.K. Powerscreen unveiled three new machines – the Premiertrak 300 jaw crusher, the Trakpactor 320SR impact crusher and the Warrior 2100 screen as well as showcase some well-established products.
Colin Clements, Powerscreen international sales director, says, "The open day was an outstanding success and feedback from distributors and customers has been fantastic. Economic conditions globally may be challenging, but Powerscreen is committed to enhancing the experience of all our customers, to reinforce our position as the world's leader in mobile crushing and screening equipment for our key sectors and to continue to innovate as we have in the past."
Four machines were working in recycling applications, processing tarmac, concrete, brick and mixed demolition waste:
Premiertrak 300 HA Hydrostatic VGF jaw crusher
The Premiertrak 300 jaw crusher featured the new optional hydrostatic drive system allowing it to reverse and unblock or run continuously in reverse for asphalt applications. It is configured to run at low engine RPM resulting in excellent fuel efficiency and low noise emissions, ideal for urban sites. The Premiertrak 300 crusher was configured with the Vibrating Grizzly Feeder (VGF) set with 50 millimeters (mm) bofor bar on the top deck and 30 mm mesh on the bottom. The closed side setting (CSS) was set to 40 mm while running in both asphalt and recycled concrete applications.
Other benefits of the machine include: Large clearances in the bypass chute to improve material flow and reduce the chance of blockages in sticky material; a large under crusher clearance and full tunneling to reduce the chance of blockage due to rebar build up; and a hydraulically lowering conveyor offers excellent access to reduce downtime should a blockage occur.
Rapid set-up times are possible with all folding done hydraulically and no need to work at height so the machine can be moved very quickly. Control is via an easy to use PLC screen control system. Jaw speed and CSS can be changed by a single button which allows the machine to be adjusted to suit the application very quickly with no tools required.
Premiertrak 400 Pre-Screen jaw crusher
The Powerscreen Premiertrak 400 jaw crusher has been developed to incorporate a fully independent high amplitude, hydraulically driven pre-screen with increased drop angle on the bottom deck improving machine throughput and overall capacity. In order to facilitate improved pre-screened material flow, the discharge opening from the under chute has been increased from 200 mm to 320 mm, allowing faster movement of pre-screened material to bypass the crushing chamber.
The Premiertrak 400 was fitted with pre-screen bofor bars on 60mm top deck and 30mm on the bottom deck. Machine closed side setting was 45mm and processed recycled concrete containing a high percentage of rebar.
Trakpactor 320SR impact crusher
The Trakpactor 320SR is a mid-sized horizontal impact crusher redesigned with some key enhancements to offer operators and contractors excellent reduction and high consistency of product shape for performance in quarry and recycling applications. The plant is capable of working in the most demanding environments and features rapid set-up time, fuel efficient direct drive system and excellent output potential of up to 352 tons per hour (tph) (320 metric tph). The plant features a recirculating conveyor which raises for transport allowing more ground clearance under the machine when moving on site and loading for transport. One of the most exciting features of the Trakpactor 320SR crusher is the quick-detach post-screen section which converts it to a standard Trakpactor 320 unit. This provides operators with outstanding versatility to use the machine in a variety of different applications.
Warrior 2100 screen
The Warrior 2100 screen has been engineered to include the proven Triple Shaft technology first developed for the successful Warrior 2400, a model which is now well established in markets worldwide. This Triple Shaft technology, unique to Powerscreen in heavy duty mobile screens, is designed so the 16' x 5' screen box is highly effective and efficient while maintaining exceptional throughput productivity. The extreme screening acceleration offers the Warrior 2100 improved capabilities over its class rivals, especially in sticky scalping applications. During the event, the machine processed mixed demolition waste which included grass, soil, concrete, wood and asphalt, and was configured with a combined top deck of 60mm fingers with 60mm punch plate and a bottom deck of 40mm mesh.
Four machines were on static display:
Trakpactor 250 impact crusher
Designed especially for the recycling and demolition markets, the Trakpactor 250 impact crusher has special features designed to prevent chamber overload and damage. Capable of processing a wide range of material at a high output, the plant is versatile, easy to operate and fuel efficient due to its direct drive system.
1000 Maxtrak cone crusher
The high performance Powerscreen 1000 Maxtrak is a small to medium sized cone crusher which has been designed for direct feed applications without pre-screening on clean rock. At its heart is the Automax cone crusher which provides excellent reduction and product cubicity for the production of high quality aggregate and sub-base material. Hydraulic release capabilities are designed so that any uncrushable material entering the chamber is safely discharged.
Chieftain 1700 screen
The Powerscreen Chieftain 1700 is a mid-sized track mobile screen aimed at end users requiring high volumes of finished products in applications including topsoil, coal, crushed stone, recycling, iron ore, sand and gravel.
Warrior 1400X screen
The 1400X has been designed with economy in mind, with reduced engine running speed and enhanced hydraulics, the 1400X offers a reduction in fuel consumption of up to 15 percent in comparison to its predecessor. This can translate into a substantial financial saving for customers over the lifetime of the machine.
Visit us at ConExpo 2014, Terex booth 1047 on the Gold Lot, to see the Premiertrak 300, Trakpactor 320SR and Warrior 2100 units on display.
For more information on Powerscreen crushing and screening products or customer support, please contact your local dealer. Details can be found at www.powerscreen.com.
Genesis Attachments adds vice president-general manager: Jeff Adams was previous... Wednesday, April 01, 2015 05:31:12 AM
NorthStar Demolition begins dismantling Daytona track's Sprint Tower: Month-long... Wednesday, April 01, 2015 04:27:06 AM
Sacramento Kings demo project tops recycling goals: Demolition tied to new arena... Wednesday, April 01, 2015 03:48:08 AM
Baetsen Recycling adds Sennebogen electric handler: Netherlands recycler is usin... Tuesday, March 31, 2015 04:27:39 AM
Don't miss another issue. Subscribe to Construction & Demolition Recycling now
w... Saturday, March 28, 2015 11:30:35 AM
Subscribe to the magazine you love to follow today at www.cdrecycler.com/subscri... Friday, March 27, 2015 01:49:06 PM
Eriez offers equipment service case studies http://dlvr.it/97zNdm Friday, March 27, 2015 11:36:40 AM
REM offers new portable sorting system http://dlvr.it/97MTf4 Thursday, March 26, 2015 03:51:27 PM
Genesis adds new shears to XT Mobile line: New GXT models offered in straight an... Thursday, March 26, 2015 07:19:10 AM
Waste symposium issues call for abstracts: Global Waste Management Symposium wil... Wednesday, March 25, 2015 01:42:07 PM
Bandit offers new options for horizontal grinders: Chipper manufacturer's Beast... Wednesday, March 25, 2015 01:30:10 PM
Masaba introduces new portable crusher http://dlvr.it/96PWsh Wednesday, March 25, 2015 10:27:41 AM
City of Phoenix seeks landfill diversion ideas: The city's "call for innovators"... Tuesday, March 24, 2015 08:54:39 AM
Superfund reports now available through ISRI bookstore: The reports are designed... Tuesday, March 24, 2015 04:04:40 AM
ClearSpan Fabric Structures introduces Giant Doors: Entryways can be custom desi... Monday, March 23, 2015 04:03:36 PM
ClearSpan Fabric Structures introduces Giant Doors http://dlvr.it/94xQcg Monday, March 23, 2015 03:45:38 PM
Eco Expo Asia set for Oct. 28-31, 2015 http://dlvr.it/94wvQ9 Monday, March 23, 2015 03:27:09 PM
Sennebogen adds Anderson Equipment as dealer http://dlvr.it/934DKT Friday, March 20, 2015 05:55:11 PM
General Kinematics appoints new president http://dlvr.it/92J6bh Thursday, March 19, 2015 04:53:14 PM
Think tank questions New York brownfields plan: Budget scrutiny group says Empir... Thursday, March 19, 2015 04:42:12 PM
Neighbors greet Houston demolition with enthusiasm: Residents happy to see empty... Thursday, March 19, 2015 04:25:13 PM
Federal officials sue Maine demolition firm over unpaid fines: Lawsuit seeks to... Wednesday, March 18, 2015 03:35:09 PM
Majority of US experiences construction job growth in the last year: AGC analyst... Wednesday, March 18, 2015 03:29:15 PM
Republic Services cites recycling, renewable energy in Sustainability Report: Mo... Wednesday, March 18, 2015 11:53:10 AM
The March/April issue of Construction & Demolition Recycling is now available:... Tuesday, March 17, 2015 12:34:44 PM
CDRA announces C&D recycling industry award winners: Winners will be recognized... Tuesday, March 17, 2015 10:55:16 AM
Hartl introduces two-deck screening plant: Austrian-built HSP 3300 is used to si... Tuesday, March 17, 2015 05:34:41 AM
DustBoss Mini designed for precise placement http://dlvr.it/8zvhxp Monday, March 16, 2015 02:03:47 PM
ISRI releases programming details for 2015 convention: Sessions will cover a var... Monday, March 16, 2015 06:07:42 AM
Metso mobile preshredders are designed for easy transport http://dlvr.it/8xrmsg Friday, March 13, 2015 09:28:14 AM
5811 Canal Road
Valley View, OH 44125 USA


text:
Supported URL Shortener:
* is.gd
* goo.gl (with and without Authentication)
* bit.ly / j.mp (with and without API Key)
* sck.pm
* tinyurl.com
* to.ly
* ur.ly
* urlr.eu
* yourls (own installation with signature)
If you find any Bugs, please report them to the Bugtracking at http://issues.keineantwort.de/.
If you are missing your language, please submit your translation at http://www.getlocalization.com/URLShortener/.
If you have any problems, questions or ideas, mail me at firstname.lastname@example.com
Tags: URL, Shortener, Share URL, twitter, tweetdeck, tweet
What's New
Open Shared URL
Copy URL
For example, pressing the share button in the YouTube app and selecting 'Copy URL' will copy the videos URL to the clipboard and disregard extra text like the title information.
*This app has no GUI or icon. It is only visible in share menus and Settings->Apps*
How To Use:
-Open an app that uses a share menu
-In the share menu, choose Copy URL
-If a valid URL is found, it will be copied to the clipboard
-Paste the copied URL wherever you would like
If you like this app, please consider purchasing the pro/donate version.
Any Questions?
Please contact me!
ShortLink shortens your links and returns a URL which have been shortened by one of it's supported services. It also generates a QR code which you can share with your friends if need be. With this app you can easily pass links/URL's from other apps into this app, shorten in, and quickly share them through other apps such as facebook, twitter, email etc.
Stores the shortened link history on your phone for easy access. If need be, it can also be cleared.
Currently supports the following providers:
* goo.gl
* bit.ly
* tinyurl
* turl.ca
* ow.ly
* is.gd
Do you miss a provider? Let me know and I'll see what i can do.
Coming soon:
* Log in to the providers which supports it
* Click statistics
* Loads of other nifty stuff
URL Notification
This app can record URL sent to notification panel. Recent 100 entries are saved. To use it, you need to enable this function from settings.
This app is able to work as plug-in for twicca (Twitter client). In the situation that you are reading your timeline, you can send URL in a tweet to notification panel quickly and open the URL after finished reading timeline.
This app accesses to the Internet to get the page title.
[How to use]
1. Select "Share page" menu in a web browser or other app
2. Select this app
3. The URL is sent to notification panel
Browser Intercept - Share URL
Note: The option to share URL will disappear if you select a default browser. The app is only useful for those who want to choose something different every time they open a URL on their device.
Url Shortcut
NOTE: This app does NOT have its own launcher icon. The only way to "open" it is to long-press a blank area of your home screen and choose to add a shortcut using 'Url Shortcut'. Some new launches hide shortcuts amongst the other widgets. If your launcher has a widgets drawer then you should look in there for a 1x1 Url Shortcut widget.
You should use a good launcher like ADW Launcher, Launcher Pro or Go Launcher which lets your specify custom icons. This means that I don't have to bundle a billion icons with this app.
NOTE 2: Use 'URL Shortcut' and the 'Desktop VisualizeR' to create shortcuts with whatever icons you like:
If you give this app 1 star, you have anger management issues and expect too much from the people around you. You will be unable to find someone to care for you and this will benefit the rest of us by stopping your negative genes from propagating to the rest of the population. If you have problems (with the app, not in general) or suggestions, press the contact button in the Android market app :)
People with anger management issues:
Jeffzda - you've missed the point of this app. The idea is to NOT clog up your browser's bookmarks with more URLs.
Byron - There's no misrouting going on, that's Android not knowing what to do with a mailto: URL. The shortcut this app creates simply has 'data' set to whatever you give it. If I claimed to treat mailto: URLs any different then fair enough, but I didn't. To the anger clinic with you!
Lisa - Lisa, Lisa, Lisa... did you even bother to read the application's description? There is no launcher icon for it, because that isn't how you used it. Do you also know that I can't actually reply to you to help you? Ah well, that's another 1 star attached to this app forever. You've probably uninstalled it already and forgotten about its existence. The Play Store isn't going to forget about that 1 star though :'(
Easy URL Shortener
Main features:
- Shorten URLs
- Generate QR code
- Auto-paste URL from clipboard on application start up
- Auto-copy the result URL to the clipboard
- Monitor clipboard
- Show application in status bar for faster access
- Watch history of your actions
Supported services:
- is.gd
- goo.gl
- tinyurl.com
- v.gd
- x.co
- su.pr
Feel free to contact us if you have any ideas on how to improve the application.
Short URL Resolver
Why would you need a (big) web browser to open link that sends you to a Youtube video. Click this youtube link on your Android phone: http://bit.ly/10xGRnq (Then try again after installing this app to see the difference).
Sure, there are other apps that do this, but I think mine has unique features:
* Doesn't get in the way: It's small (~700kb). Minimal, but useful, user interface. Does exactly 1 thing and does it right.
* Handles Facebook's external links. Tired of launching the browser to open a m.facebook.com?l.php URL? No more.
* Handles Youtube's attribution links. These should be handled by the Youtube app (Google's fault). Short URL Resolver handles them for you, so you get to see the video on the Youtube app. No browser involved.
* Handles Tapatalk api click links.
* Handles google.com/url links, used in the Play Store for links in the description.
* Partial support for adf.ly.
Also, it's free, and ad-free. You can make an in-app donation if you feel it's worth it.
It doesn't use any third party service for resolving the URLs. The basic principle is making an HTTP request and determining where it's being redirected.
It's my first public Android App, but I think it's the best of its kind (you can try and prove me wrong X-D)
*Thanks for the comments and the reviews!*
If anyone wants to collaborate by translating the app to another language, you can do it here: http://www.getlocalization.com/shorturlresolver/
XDA thread: http://forum.xda-developers.com/showthread.php?p=49590835
URL Monitor
Forked version of "Host Monitor"
-Allows for an optional PATH where the program will do a HTTP HEAD instead of a ping
-Fixes crashes and works in Cupcake 1.5
-New desktop widget summarizes overall status every 15 minutes (long press home screen to add)
URL File Handler
Very similar to StuntzUrl, except that this application works in any application that supports "Open With" - like RootExplorer and GooglePlay.
ShortPaste - Autoshorten URLs
Tired of sharing long, nasty URLs? ShortPaste immediately shortens URLs after copying them (from an app such as Chrome, YouTube, Newsstand, etc) in the blink of an eye. Like magic, your long URL will appear in its shortened form when pasted. Supports bit.ly and goo.gl.
Follow the development on XDA: http://bit.ly/shortpastexda
Above Android: http://goo.gl/dihQD2
URL from QRCode to browser
It opens internet browser immediately after QR code detection.
Point camera to code and get site opened!
Use history page to open any site again.
Best together with "QR-Code Tag Extension" plugin for Google Chrome on your PC.
Tasker URL Launcher
Useful for launching tasker tasks from NFC tags-
for Sony Xperia smarttags simply use the the smart tag app. To run a tasker task use the "Open URL" action, then set the url "tasker://taskname" instead of "http://" (change taskname to the name of your task defined in tasker- tasks are case sensitive)
e.g. "tasker://mytask" to run a task called "mytask" defined in tasker.
for writable tags it should work as simply as writing the tasker:// url and task to the tag itself.
It will do until tasker add nfc capability :)
On some devices the "tags" app comes pre-installed and blocks the url from nfc tags from being launched automagically, meaning you need to touch the screen to launch the task. You may find disabling the "tags" app from settings->apps->all fixes this problem.
Also it is VITAL that tasker is installed before this app or it will not work, this means that if you upgrade or reset your device etc. and google play automatically re-installs all your apps then it may not work. this is solved by simply uninstalling tasker url launcher, checking tasker is installed, and then re-installing the tasker url launcher from google play.
URL Shortener - make money
When you copy some URL it will appear in its shortened form when pasted, automatically!
How to:
- Go to https://shorte.st/ref/aebff85382
- Register or Log in
- Go to https://shorte.st/tools/api
- Copy your API Token
- Paste it in application settings
- enjoy!
- automatically shorten links that you have copied
- displays notification when link is copied with ability to shorten it or dismiss
- keeps history of your shortened links
- ability to share link with almost every possible way
- generate QR code with shortened link
- share generated QR code
This is not official app!
URL Scheme Sender
You will be able to easily send the Intent using URLScheme.
You can save 30 history.
key words
URLSchemeSender, Manifest, Test
URL Shortener
Shorten long URLs and squeeze them into fewer characters to make links easier to share, tweet, or email to friends. Using a shortened goo.gl URL leaves more room to say what's on your mind. For example, http://goo.gl/n7IMF2 is a convenient shorthand representation for the long link https://play.google.com/store/apps.
Access analytics of any short URL, including the raw click counts and their distribution across referrers, browsers, platforms, and geographical locations.
The app also lets you
• Shorten URLs and share them quickly
• Access your history - always synced with goo.gl
• Get colorful analytics reports of any short URL, not just yours
• Android Beam - push a short URL onto another device by tapping the devices together
• Shorten links through "share" in any app and get rich notifications
• Star your favorites
• Generate and share QR codes
Have a bug report or feature request? Please contact me by email or on Google+.
You can help translating this app! Contact email@example.com for more information.
Thanks infinitely to all who already contributed: extendwings, BrunoMioto, fedec96, siveZ, johan98, Bigomby, HeinrichReimer, SHURman, keem1, Yari27, echelonx, Froony, aniquewetzels and many others.
This app is in no way affiliated nor endorsed by Google Inc.
More from developer
Ist mein Zug pünktlich?
m.bahn.de bietet die Funktion "Ist mein Zug pünktlich?".
Mit dieser App kannst Du oft gefahrene Züge speichern und so schnell und einfach auf "Ist mein Zug pünktlich?" zugreifen.
Seit neuestem werden auch die Metronomzüge durch die Bahn-Website unterstützt. Um die Funktion "Ist mein Zug pünktlich?" für einen Metronom (z.B. ME82003) zu nutzen einfach "RE/RB" als Zugtyp auswählen und als Zugnummer die Nummer eingeben (z.B. 82003). Der Zugtyp "ME" fällt weg.
S-Bahn wird teilweise von m.bahn.de nicht unterstützt, daher kann ich es auch nicht unterstützen. Wenn mir jemand die Möglichkeit bringt, dass ich an die Daten komme, kann ich auch Nahverkehr unterstützen. Der HVV bietet keine Möglichkeit an die Daten zu kommen.
Weiterhin kann man eine Haltestellen-Abfrage speichern und so An- und Abfahrten an einer Haltestelle sehen. Hier werden auch andere Verkehrsmittel unterstützt (bspw. S-Bahn, Bus, Fähre...)
Sollte Deine Haltestelle nicht gefunden werden, bitte einmal unter m.bahn.de => Ankunft/Abfahrt testen, ob die Haltestelle dort gefunden wird.
ACHTUNG: Die Daten kommen von der Bahn. Sowohl Bahnhöfe, als auch Zeiten zur Verspätung kommen von der Bahn. Ich kann nichts dafür, wenn die Bahn falsche Verspätungen anzeigt.
Mit dem Support-Key kann die Werbung dauerhaft ausgeblendet werden. http://auchich.de/imzpp
INTERNET: Die Internet-Berechtigung wird benötigt um die Daten der Verspätungen zu laden.
SPEICHER: Die neue Backup-Funktion schreibt die Daten auf die SD-Karte
Sollte es Probleme geben, bitte einen Fehler im Issue-Management (http://issues.keineantwort.de) eintragen oder per Mail (email@example.com) / Twitter (http://twitter.com/keineantwort) melden. Eine schlechte Bewertung hilft weder Dir, noch mir.
Tags: Bahn, Deutsche Bahn, IC, ICE, Verspätung, Haltestelle, Bahnhof, Metronom


text:
Tulsa Garden Center at Woodward Park
2435 S Peoria Ave
Tulsa, OK 74114
Located in historic Woodward Park, the Tulsa Garden Center is dedicated to bringing horticultural and environmental ... read more...
14613 N Kelley Ave
Oklahoma City, OK 73013
Oklahoma City ice skating rink with pro shop and concession stand. Skating lessons and rentals available as well as ... read more...
Oklahoma City Community College Theatre
7777 S May Ave
Oklahoma City, OK 73159
The Oklahoma City Community College Theatre program's Bruce Owen Theater is a 285-seat theater located on the campus of ... read more...
Sherwin Miller Museum of Jewish Art
2021 E 71st St S
Tulsa, OK 74136
The Sherwin Miller Museum of Jewish Art educates visitors about the Jewish experience in Oklahoma, the United States, ... read more...
6 Santa Fe Plaza
Oklahoma City, OK 73102
The Red Earth Museum is a notable small museum featuring splendid examples of traditional and contemporary Indian ... read more...
Parkhurst Ranch & Pumpkin Patch
720 Henney Rd
Arcadia, OK 73007
Make your way to Parkhurst Ranch and Pumpkin Patch in Arcadia to enjoy a day of fun at a working ranch. Explore ... read more...
11521 N MacArthur
Oklahoma City, OK 73162
Strebel Creek Vineyard, Oklahoma City's first winery, was voted "Best of the Best" in 2010. Make your way to ... read more...
Myriad Botanical Gardens & Crystal Bridge Tropical Conservatory
301 W Reno
Oklahoma City, OK 73102
A lush paradise within the urban landscape, the Myriad Botanical Gardens & Crystal Bridge Tropical Conservatory is a ... read more...
The Outlet Shoppes at Oklahoma City
7624 W Reno Ave
Oklahoma City, OK 73127
Take a trip to The Outlet Shoppes at Oklahoma City for a day of shopping and good deals. The Outlet Shoppes ... read more...
Henthorne Performing Arts Center
4825 S Quaker Ave
Tulsa, OK 74105
The Henthorne Performing Arts Center, operated by the city of Tulsa, houses both the Heller Theatre and the Clark ... read more...
Email: email@example.com


text:
Tatty Teddy
engelbtcJuly 14, 2006
I live in South Africa. I found Tatty Ted recently and think it is the cutest cross stitch. Not all the kits are available in South Africa as yet. Those i could find are very expensive. Does anyone have just the charts to share? I can't find any charts that are not in the kits. Please let me know if you have any to share. I want to make some of the tatty teds for my best friends baby to be born in later this year. You can email them to me at email@example.com. Much appreciated
Dear Claire,
Dear Claire,
try to acess: http://janainarogerio.multiply.com/photos/
I am sure you will find something ...
Get in touch!


text:
Therapists: 1 to 20
Refine Your Search
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Treatment Orientation:
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Broaden Your Search
See Nearest
Therapists in Mount Airy - Philadelphia, PA 19119
Verified by Psychology Today
Elizabeth Pollak
Clinical Social Work/Therapist, MA, LCSW
"ARE YOU CONCERNED ABOUT YOUR CHILD'S WELL-BEING? Is your teen challenging you in ways that feel overwhelming? I welcome your child to work through areas of concern to him/herself and others. I provide support to children dealing with mood changes; school difficulties and stressful relationships. I include family members in this process, unless circumstances preclude this. I utilize play therapy and other therapeutic interventions. IN ADDITION, I SEE INDIVIDUAL ADULTS, who are confronting life transitions; mood changes; relationship concerns and histories of trauma, among other sources of emotional pain. I can be reached at 215 687 7570.
(267) 396-1948
Verified by Psychology Today
Lisa Mullins Marchiano
Clinical Social Work/Therapist, LCSW
"Deciding to speak to a stranger about your personal life can be daunting. Finding the right therapist is often a matter of "fit." I don't believe that I know what is right for you. I believe something in you knows what is right for you, and that it is my job to help us listen for that inner knowing. I have an interactive style that is challenging and compassionate. The nature of my work with clients is determined first and foremost by your needs and preferences.
(267) 348-4078
Verified by Psychology Today
Jane K Shure
Clinical Social Work/Therapist
"I'm a national expert on bolstering resilience and self-confidence, helping people identify strengths, manage emotions, overcome resistance to change and mobilize forward movement. Named a Philadelphia Magazine "Top Doc for Women," I'm a published author on issues of shame, trauma, body-image, eating disorders. I lead workshops on Calm Your Inner Critic & Strengthen Your Inner Coach, am on faculty at Kripalu Center and Athena Leadership Lab, coauthored a teen self-esteem building curriculum, and coedited Effective Clinical Practice in the Treatment of Eating Disorders. For more see: huffingtonpost.com/jane-shure/, SelfMatters.org and resilientleadership.org/blog.
(215) 995-1198
Verified by Psychology Today
Tracy Weant
Clinical Social Work/Therapist, MSW, LCSW
"Everyone deserves a safe space to be heard, seen and supported. It is my intention to create such a space. I understand that life is messy and I will work with you to solve life's challenges as they arise.
(267) 973-8233
Verified by Psychology Today
Ameet Ravital
Psychologist, PhD, DiHom
"Are you experiencing emotional distress, relationship challenges, financial stress or chronic health issues? Over time, such problems can be exhausting to your mind, body and spirit. Unresolved, they can take a heavy toll on your work performance and daily satisfaction with life. I can offer you a very different approach. Change will start with the first session. Together we'll start by understanding your goals, and then we'll lay out the first steps to move in that direction. Aren't you ready to uncover new possibilities?
(215) 987-5340
Verified by Psychology Today
Sondra Rosenberg
Art Therapist, ATR-BC
"My approach as an art therapist is humanistic and image-based. I help clients to find projects that best enable them to express their inner world. Together, we enter into that world and explore the contents therein. This exploration opens up possibilities and solutions in areas where the client is stuck. I believe that each element of an image reflects different parts of the person who created it. Through the use of voice dialogue, where the client is guided through a process of giving voice to elements within the image, insight and self-compassion are increased and places of inner strength and wisdom are accessed.
(215) 977-4974
Verified by Psychology Today
Berth Bartolin
Clinical Social Work/Therapist, MA, LPC
"Life is a process which will continue to present many challenges and enjoyable events. We will work together to build your positive coping skills while focusing on your confidence and overall wellness. I am here to help you discover your identity and strengths. You will grow to continue to set your place in the community.
(267) 433-0065
Verified by Psychology Today
Cassandra Bolding
Counselor, LPC, CAC, ICADC, HIVspec
"Provision of counseling/therapy/coaching to adolecesant through adult.Specialties include addictions(drug/alcohol, gambling, shopping, sex, food),womens issues,and post traumatic stress disorder resulting from(violence, combat, sexual trauma). In addition to HIV/AIDS issues,newly diagnosed and partners welcomed. Types of services offered include couples, family,group and individual therapy/coaching. We will work collaboratively to address depression, anger, fear, grief/loss, stress, relationship concerns and anxiety that impair your quality of life.
(215) 240-8617
Verified by Psychology Today
Carrie Askin
Clinical Social Work/Therapist, LCSW
"Do you have difficulty connecting to your partner or maintaining long-term, meaningful friendships? Are you often lonely and wishing for connection? Do you want to eliminate drama in your relationships? Therapy can help you to create more meaningful relationships with yourself and with others. In therapy, we work to develop a clarity about your needs and desires and to build the skills you need to authentically engage with others, to develop comfort with conflict and to learn to stay connected with ourselves and others even as we express our differences clearly.
(267) 613-4305
Verified by Psychology Today
Rebecca K Stern
Psychologist, PsyD, PhD
"I believe a good match between therapist and client is a large part of what makes therapy work. I have a caring, gentle approach and I take a mind-body approach to psychological issues. I accept Medicare, Aetna, and Bluecross/ Blueshield insurances. I also work with children on stress management and social anxiety issues, and received training in family therapy.
(919) 295-2438
"If you are having difficulty overcoming struggles in your life, you are not alone. I can help you navigate through these issues, and offer a safe, comfortable, confidential space for you to share. I listen with empathy and can assist you in defining appropriate strategies to help you meet your treatment goals. My clientele consists of children, adolescents, and adults; I have experience working with a variety of issues including: ADHD, ADD, Autism, Anxiety, Depression, Anger, Adjustment issues, low self-esteem, poor coping skills, and interpersonal relationship problems.
(267) 723-7534
Daniel Silverman
Counselor, MEd, JD
"I am not beholden to any one counseling model. What is most important to the therapeutic process is the contact we share in our sessions and my ability to create a safe environment for you to explore and work on yourself and your relationships. I facilitate that process. I do both individual and couples counseling. I am most interested in working with couples who wish to make their relationships deeper and more gratifying and with high-functioning individuals who have the basics covered but are searching for greater satisfaction in life.
(215) 240-1876
Verified by Psychology Today
Robin Frederick Witts
Clinical Social Work/Therapist, MSW, CS, CPRP
"I have been working in community mental health for several years providing psychotherapy, supportive counseling and grief counseling. I believe in the strength of the human spirit and I view the client as the expert in their journey to wellness. I hold strength in the statement "I can walk with you but I cannot walk for you".
(267) 354-7308
Verified by Psychology Today
Amelia Klein
Clinical Social Work/Therapist, DSW, LMFT, LCSW, BCD
"At this moment, you may be involved in a relationship in need of repair, dealing with issues from your past, or grieving the loss of a loved one. You may be feeling lost or without direction. Maybe you're self-medicating by excessively using drugs and/or alcohol. You may be feeling unhappy in your relationships or just about to make an important life transition. Whatever may be troubling you, I believe I can help you.
(215) 247-2819
Verified by Psychology Today
Genie Bud Ravital
Clinical Social Work/Therapist, MSS, LCSW
"Change is possible! It thrives in a warm, supportive space that allows personal introspection and gentle encouragement. I have helped people with a wide range of problems, including relationship conflicts, parenting issues, chronic depression, dealing with traumatic events, and substance abuse. I think of myself as a practical kind of therapist. I predominantly use cognitive behavioral therapy to help you change negative thinking patterns and develop healthier choices in taking care of yourself and better communication in your relationships. I can help you identify the problems you want to solve and the specific goals you want to achieve.
(267) 436-7178
Verified by Psychology Today
Jennifer Ellis
Counselor, MEd, LPC
"I am a licensed professional counselor with several years of experience, treating adults in a large variety of contexts. I provide both individual and couples counseling, specializing in treatment of anxiety disorders, mood disorders, and psychotic disorders. The treatment I provide is client-centered, using a cognitive-behavioral approach (CBT), which is solution-focused and geared toward lessening distress that can result from life struggles, self-esteem, symptoms, and relationship issues. Because the decision to enter therapy can often be very difficult, the foundation of the treatment that I provide starts with our relationship.
(609) 879-4454
Verified by Psychology Today
Harin Feibish
Clinical Social Work/Therapist, MA, LCSW
"With 30+ years of experience, I enable clients to become aware of how their thoughts and behaviors inhibit daily life, acknowledge how this effects them and others, and take action to move forward. I am a nationally recognized authority on the treatment and prevention of eating and body image disorders. I founded and formerly directed "Body Balance" at The Renfrew Center, a program which focuses on women's eating and body image issues. My work has included numerous groups, workshops, written articles, and TV appearances that have focused on issues of self-acceptance and self-esteem.
(215) 789-9423
Verified by Psychology Today
Eric Robert Mitchell
Psychologist, PhD
"I predominately specialize with autistic clients and their families, having authored the "The Autism and Asperger's Handbook: Putting It Together" which is available on iTunes/iBookstore. I also enjoy a variety of therapeutic and consulting opportunities, including working with schools, service providers, self-advocates, couples, families. I also enjoy connecting families with systems such as schools, community resources, services, and so on, facilitating family navigation through difficult times. I tend to specialize with families, children, and young adults affected by autism, although practice a wide array of interventions for many populations.
(215) 839-1218
Verified by Psychology Today
Michelle Keyes
Clinical Social Work/Therapist, MA, LPC
"Michelle is a licensed psychotherapist with over 10 years experience working in the field of mental health. In addition to her general practice, Michelle has a special interest in the emotional wellness of women and provides therapy to meet the unique needs of women with depression, anxiety, chronic stress and other mental health problems. Michelle understands that life experiences such as relationship changes, parenting, reproductive transitions (pregnancy, postpartum, menopause), and issues of grief and loss can influence a woman's vulnerability to emotional disturbances. With complementary therapy, women can begin overcoming these challenges and experience a life of emotional wellness.
(215) 874-6177
Verified by Psychology Today
Anne D Ream
Counselor, MFT, LPC, ATR-BC
"It is your right to high quality affordable counseling. With 25 years experience in counseling and 20 years experience in developmental psychology, I can give you that quality. Are you struggling with anxiety or depression because of a difficult person? Is your significant relationship struggling? Have you experienced or witnessed difficult/traumatic events? Have you experienced a divorce or death of a loved one? Are you a single parent? Many life stressors result in a person needing to talk with someone with a deep understanding of people. I have the knowledge and experience to give you, as an individual, the kind of help you need.
(215) 995-5248
Not enough Therapists in Philadelphia County, Pennsylvania (PA), 19119 to choose from?
Try expanding your search for Therapists in 19119 to a larger area (e.g. click the plus sign or 19119+ .)
How can I tell if a therapist is right for me?
Therapists in Mount Airy - Philadelphia have broad skills and are able to work with a wide range of issues.
For example, if you're seeking a marriage counselor in 19119 you'll find that most therapists are trained in marriage counseling or couples counseling in 19119 and couples therapy. And they welcome families for family counseling in 19119 or family therapy in Mount Airy - Philadelphia.
How to find a specialist?
If you're looking for specialist counseling in Philadelphia County, Pennsylvania 19119, such as a child psychologist or sex therapist, try to refine your search with our left nav bar.
What method of therapy is right for me?
When it comes to treatment methods, there's no "right way". A couples counselor or a family counselor may use different treatment methods depending on the one that works best for you.
How to use my insurance?
Many therapists accept health insurance. Check to see if your insurance is covered. Feel free to discuss this when you contact the therapist.


text:
|Reviews for Flame Jockeys Episode 1
| Omne chapter 1 . 4/9/2011
Hey you know what? I think you could make it as a TV writer. I mean this is very good writing and I think it would make a good show. Oh if only someone would produce it for you. Anyway, me thinks this is very very good and it's too bad you can't turn it into a real TV show.
| JOE chapter 1 . 10/21/2005
hi-the story was good-i liked it was written in screen-play format,it was easy for me to was kool cause you got me at the end.i had no idea that austin was going to end up fooling around with mack. the dream sequence threw me off. its obvious she has strong feelings for merrill. bob and brad hint at merrill s feelings for austin at the end. so, leaves me hanging-thats-good-lol-to be critical wouldn t be fair-i have to read the other episodes-keep writin:)
| ArchDemonNotion chapter 1 . 10/17/2005
Very very good script writing. It's funny as all hell and very entertaining. I admit I had some LOL moments.
| dragonelemental chapter 1 . 6/10/2005
A few criticisms but none of them very big... Again, i loved it. very funny, i loved the relationship that the guys showed they had at the beginning and the opening drunk scene was funny too, catches the audience. I assume it's supsed to be a script for a television series? I think it would need to be a bit longer if u wanted to do anything with it, have you thought of actually sending it away to a producer and seeing what they think? You've got all of the essentials, character development and relationships, main plots and themes, it could go all the way. Again the fact that it was in script form put me off at firt but when i understood what it was i thought it was bloody amazing. Really cool. Ok, i'm gonna go look at more...