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https://www.readbyqxmd.com/read/28815506/preparing-fosmid-mate-paired-libraries-using-cre-loxp-recombination
#1
Ze Peng, Jeff L Froula, Jan-Fang Cheng
Fosmid end sequencing has been widely utilized in genome sequence assemblies and genome structural variation studies. We have developed a new approach to construct fosmid paired-end libraries that is suitable for Illumina sequencing platform. This approach employs a newly modified fosmid vector (pFosClip) which contains two loxP sites with identical orientation and two inverse Illumina adaptor priming sites flanking the cloning site. DNA prepared from the fosmid library constructed with pFosClip can be treated with the Cre recombinase to remove most of the vector DNA, leaving only 107 bp of the vector sequence with insert DNA...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28814585/a-multiplex-rt-pcr-assay-for-the-simultaneous-identification-of-neisseria-gonorrhoeae-and-its-ciprofloxacin-susceptibility-status
#2
Sumudu R Perera, Nurul H Khan, Irene Martin, Ali Taheri, Rajinder P Parti, Paul N Levett, Greg B Horsman, Anthony Kusalik, Jo-Anne R Dillon
A real time PCR (RT-PCR) assay was designed for the simultaneous identification of N. gonorrhoeae and its ciprofloxacin susceptibility status. A SYBR-Green based multiplex RT-PCR format was used and comprised two different forward primers and a common reverse primer to detect single nucleotide polymorphisms (SNPs) in gyrA of N. gonorrhoeae The primer pairs were evaluated for their sensitivity and specificity with genomic DNA from 254 N. gonorrhoeae isolates (82 were ciprofloxacin susceptible and 172 were ciprofloxacin resistant) and 23 non-N...
August 16, 2017: Journal of Clinical Microbiology
https://www.readbyqxmd.com/read/28814526/hiv-1-exploits-dynamic-multi-aminoacyl-trna-synthetase-complex-to-enhance-viral-replication
#3
Alice A Duchon, Corine St-Gelais, Nathan Titkemeier, Joshua Hatterschide, Li Wu, Karin Musier-Forsyth
A hallmark of retroviruses such as human immunodeficiency virus type 1 (HIV-1) is reverse transcription of genomic RNA to DNA, a process that is primed by cellular tRNAs. HIV-1 recruits human tRNA(Lys3) to serve as the reverse transcription primer via an interaction between lysyl-tRNA synthetase (LysRS) and the HIV-1 Gag polyprotein. LysRS is normally sequestered in a multi-aminoacyl-tRNA synthetase complex (MSC). Previous studies demonstrated that components of the MSC can be mobilized in response to certain cellular stimuli, but how LysRS is redirected from the MSC to viral particles for packaging is unknown...
August 16, 2017: Journal of Virology
https://www.readbyqxmd.com/read/28813779/dna-profiling-of-leuconostoc-citreum-strains-in-fermented-foods-by-repetitive-element-polymerase-chain-reaction
#4
Jasmine Kaur, Anshul Sharma, Sulhee Lee, Young-Seo Park
To identify and discriminate the bacterial species at the sub-specific level, rep-PCR method is a reliable genomic fingerprinting tool. Fourteen strains of bacteria were isolated from different food sources, identified as Leuconostoc citreum using 16S rRNA gene sequencing and amplified using rep-primers (REP, ERIC, and (GTG)5). Fingerprinting patterns generated band range in between 300-6000 bp with REP, 150-6000bp with ERIC and 200-1700 bp with (GTG)5 primers respectively. In UPGMA dendrogram analysis, 14 strains were clustered into three clades (I, II and III) with all the primers, thus differentiating them at the molecular level...
August 17, 2017: Journal of Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28813520/the-frequency-of-class1-and-2-integrons-in-pseudomonas-aeruginosa-strains-isolated-from-burn-patients-in-a-burn-center-of-ahvaz-iran
#5
Azar Dokht Khosravi, Moloudsadat Motahar, Effat Abbasi Montazeri
BACKGROUND: Pseudomonas aeruginosa is an opportunistic pathogen with the ability to cause severe nosocomial infections and remains a major problem in burn patients. This organism shows a remarkable antimicrobial resistance and is often resistant to multiple antibiotics. Integron genes as mobile genetic elements are playing an important role in the spread of P. aeruginosa antibiotic resistance. This study was aimed to investigate the occurrence of class 1, and 2 integron genes (int1, int2), among P...
2017: PloS One
https://www.readbyqxmd.com/read/28812169/bond-strength-of-etch-and-rinse-and-self-etch-adhesive-systems-to-enamel-and-dentin-irradiated-with-a-novel-co2-9-3%C3%A2-%C3%AE-m-short-pulsed-laser-for-dental-restorative-procedures
#6
Peter Rechmann, N Bartolome, R Kinsel, R Vaderhobli, B M T Rechmann
The objective of this study was to evaluate the influence of CO2 9.3 μm short-pulsed laser irradiation on the shear bond strength of composite resin to enamel and dentin. Two hundred enamel and 210 dentin samples were irradiated with a 9.3 µm carbon dioxide laser (Solea, Convergent Dental, Inc., Natick, MA) with energies which either enhanced caries resistance or were effective for ablation. OptiBond Solo Plus [OptiBondTE] (Kerr Corporation, Orange, CA) and Peak Universal Bond light-cured adhesive [PeakTE] (Ultradent Products, South Jordan, UT) were used...
August 15, 2017: Lasers in Medical Science
https://www.readbyqxmd.com/read/28811759/seed-transmission-of-tomato-yellow-leaf-curl-virus-in-white-soybean-glycine-max
#7
Eui-Joon Kil, Jungho Park, Hong-Soo Choi, Chang-Seok Kim, Sukchan Lee
Tomato yellow leaf curl virus (TYLCV) infection of the common bean (Phaseolus vulgaris) has been reported, but soybean (Glycine max) has not previously been identified as a TYLCV host. Five cultivars of white soybean were agro-inoculated using an infectious TYLCV clone. At 30 days post-inoculation, they showed infection rates of 25% to 100%. Typical TYLCV symptoms were not observed in any inoculated plants. To examine whether TYLCV was transmitted in soybean seeds, DNA was isolated from bundles of five randomly selected seeds from TYLCV-inoculated soybean plants and amplified with a TYLCV-specific primer set...
August 2017: Plant Pathology Journal
https://www.readbyqxmd.com/read/28811308/wilms-tumour-1-wt1-in-development-homeostasis-and-disease
#8
REVIEW
Nicholas D Hastie
The study of genes mutated in human disease often leads to new insights into biology as well as disease mechanisms. One such gene is Wilms' tumour 1 (WT1), which plays multiple roles in development, tissue homeostasis and disease. In this Primer, I summarise how this multifaceted gene functions in various mammalian tissues and organs, including the kidney, gonads, heart and nervous system. This is followed by a discussion of our current understanding of the molecular mechanisms by which WT1 and its two major isoforms regulate these processes at the transcriptional and post-transcriptional levels...
August 15, 2017: Development
https://www.readbyqxmd.com/read/28810858/rapid-and-sensitive-detection-of-canine-distemper-virus-by-real-time-reverse-transcription-recombinase-polymerase-amplification
#9
Jianchang Wang, Jinfeng Wang, Ruiwen Li, Libing Liu, Wanzhe Yuan
BACKGROUND: Canine distemper, caused by Canine distemper virus (CDV), is a highly contagious and fatal systemic disease in free-living and captive carnivores worldwide. Recombinase polymerase amplification (RPA), as an isothermal gene amplification technique, has been explored for the molecular detection of diverse pathogens. METHODS: A real-time reverse transcription RPA (RT-RPA) assay for the detection of canine distemper virus (CDV) using primers and exo probe targeting the CDV nucleocapsid protein gene was developed...
August 15, 2017: BMC Veterinary Research
https://www.readbyqxmd.com/read/28810735/accurate-electrochemistry-analysis-of-circulating-methylated-dna-from-clinical-plasma-based-on-paired-end-tagging-and-amplifications
#10
Feng Chen, Xuyao Wang, Xiaowen Cao, Yongxi Zhao
Circulating methylated DNA has been a new kind of cancer biomarker, yet its small fraction of trace total DNA from clinical samples impairs the accurate analysis. Though fluorescence methods based on quantitative methylation specific PCR (qMSP) have been adopted routinely, yet alternative electrochemistry assay of such DNA from clinical samples remains a great challenge. Herein, we report accurate electrochemistry analysis of circulating methylated DNA from clinical plasma samples based on a paired-end tagging and amplifications strategy...
August 16, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28810700/a-single-mini-barcode-test-to-screen-for-australian-mammalian-predators-from-environmental-samples
#11
Elodie Modave, Anna J MacDonald, Stephen D Sarre
Identification of species from trace samples is now possible through the comparison of diagnostic DNA fragments against reference DNA sequence databases. DNA detection of animals from non-invasive samples, such as predator faeces (scats) that contain traces of DNA from their species of origin, has proved to be a valuable tool for the management of elusive wildlife. However, application of this approach can be limited by the availability of appropriate genetic markers. Scat DNA is often degraded, meaning that longer DNA sequences, including standard DNA barcoding markers, are difficult to recover...
August 1, 2017: GigaScience
https://www.readbyqxmd.com/read/28809012/comparative-genomics-as-a-foundation-for-evo-devo-studies-in-birds
#12
Phil Grayson, Simon Y W Sin, Timothy B Sackton, Scott V Edwards
Developmental genomics is a rapidly growing field, and high-quality genomes are a useful foundation for comparative developmental studies. A high-quality genome forms an essential reference onto which the data from numerous assays and experiments, including ChIP-seq, ATAC-seq, and RNA-seq, can be mapped. A genome also streamlines and simplifies the development of primers used to amplify putative regulatory regions for enhancer screens, cDNA probes for in situ hybridization, microRNAs (miRNAs) or short hairpin RNAs (shRNA) for RNA interference (RNAi) knockdowns, mRNAs for misexpression studies, and even guide RNAs (gRNAs) for CRISPR knockouts...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28808059/linchpin-dna-binding-residues-serve-as-go-no-go-controls-in-the-replication-factor-c-catalyzed-clamp-loading-mechanism
#13
Juan Liu, Yayan Zhou, Manju M Hingorani
DNA polymerases depend on circular sliding clamps for processive replication. Clamps must be loaded onto primer-template DNA (ptDNA) by clamp loaders that open and close clamps around ptDNA in an ATP-fueled reaction. All clamp loaders share a core structure in which five subunits form a spiral chamber that binds the clamp at its base in a twisted open form and encloses ptDNA within, while binding and hydrolyzing ATP to topologically link the clamp and ptDNA. To understand how clamp loaders perform this complex task, here we focused on conserved arginines that might play a central coordinating role in the mechanism, since they can alternately contact ptDNA or Walker B glutamate in the ATPase site, and lie close to the clamp loader-clamp binding interface...
August 14, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28807294/recombinase-polymerase-amplification-rpa-combined-with-lateral-flow-lf-strip-for-detection-of-toxoplasma-gondii-in-the-environment
#14
Y D Wu, M J Xu, Q Q Wang, C X Zhou, M Wang, X Q Zhu, D H Zhou
Toxoplasma gondii infects all warm-blooded vertebrates, resulting in a great threat to human health and significant economic loss to the livestock industry. Ingestion of infectious oocysts of T. gondii from the environment is the major source of transmission. Detection of T. gondii oocysts by existing methods is laborious, time-consuming and expensive. The objective of the present study was to develop a recombinase polymerase amplification (RPA) method combined with a lateral flow (LF) strip for detection of T...
August 30, 2017: Veterinary Parasitology
https://www.readbyqxmd.com/read/28807085/variation-in-bacterial-endosymbionts-associated-with-the-date-palm-hopper-ommatissus-lybicus-populations
#15
S Karimi, H Izadi, M Askari Seyahooei, A N Bagheri, P Khodaygan
The date palm hopper, Ommatissus lybicus, is a key pest of the date palm, which is expected to be comprised of many allopatric populations. The current study was carried out to determine bacterial endosymbiont diversity in the different populations of this pest. Ten date palm hopper populations were collected from the main date palm growing regions in Iran and an additional four samples from Pakistan, Oman, Egypt and Tunisia for detection of primary and secondary endosymbionts using polymerase chain reaction (PCR) assay with their specific primers...
August 15, 2017: Bulletin of Entomological Research
https://www.readbyqxmd.com/read/28807046/a-comprehensive-evaluation-of-the-sl1p-pipeline-for-16s-rrna-gene-sequencing-analysis
#16
Fiona J Whelan, Michael G Surette
BACKGROUND: Advances in next-generation sequencing technologies have allowed for detailed, molecular-based studies of microbial communities such as the human gut, soil, and ocean waters. Sequencing of the 16S rRNA gene, specific to prokaryotes, using universal PCR primers has become a common approach to studying the composition of these microbiota. However, the bioinformatic processing of the resulting millions of DNA sequences can be challenging, and a standardized protocol would aid in reproducible analyses...
August 14, 2017: Microbiome
https://www.readbyqxmd.com/read/28807016/endogenous-controls-of-gene-expression-in-n-methyl-n-nitrosourea-induced-t-cell-lymphoma-in-p53-deficient-mice
#17
Xi Wu, Susu Liu, Jianjun Lyu, Shuya Zhou, Yanwei Yang, Chenfei Wang, Wenda Gu, Qin Zuo, Baowen Li, Changfa Fan
BACKGROUND: Real-time polymerase chain reaction (PCR) has become an increasingly important technique for gene expression profiling because it can provide insights into complex biological and pathological processes and be used to predict disease or treatment outcomes. Although normalized data are necessary for an accurate estimation of mRNA expression levels, several pieces of evidence suggest that the expression of so-called housekeeping genes is not stable. This study aimed to validate reference genes for the normalization of real-time PCR in an N-methyl-N-nitrosourea (MNU)-induced T-cell lymphoma mouse model...
August 14, 2017: BMC Cancer
https://www.readbyqxmd.com/read/28806035/a-primer-on-diagnostic-and-financial-implications-of-d-dimer-testing
#18
Kalyan Chakravarthy Potu, Sujithasree Ketineni, Randall Lamfers
Given the high incidence of deep vein thrombosis/pulmonary embolism (DVT/PE) (300,000 to 600,000 per year in the U.S.) and the 30 percent mortality rate associated with undiagnosed PE, diagnostic evaluation is very important. One of the tools used to evaluate for DVT and PE is D-dimer testing. A negative D-dimer test, along with a low Wells clinical probability score, can safely rule out DVT/PE without the need for additional imaging. This approach can reduce cost; however, D-dimer testing is not indicated in all patients...
September 2016: South Dakota Medicine: the Journal of the South Dakota State Medical Association
https://www.readbyqxmd.com/read/28806034/a-primer-for-the-primary-care-physician-management-of-overactive-bladder-syndrome
#19
Margaret E Free, Matthew A Barker
Overactive bladder syndrome affects millions of women in the U.S. and is defined by urinary urgency, urinary frequency, and small volume voids, with or without nocturia and incontinence. Overactive bladder is a diagnosis of exclusion, and several therapies exist for the management of this condition. This article outlines a systematic approach that the primary care physician can take when treating a patient with overactive bladder.
September 2016: South Dakota Medicine: the Journal of the South Dakota State Medical Association
https://www.readbyqxmd.com/read/28806011/a-primer-on-bleeding-risk-and-management-strategies-of-newer-oral-anti-platelet-agents-in-cardiovascular-disease
#20
Jimmy Yee, Vishesh Kumar, Amornpol Anuwatworn, Marian Petrasko, Adam Stys
Aspirin, the first antiplatelet agent, has been around since the 19th century, and is one of the most established drugs in history. With the improvement of coronary interventions in the past few decades, there has been more reliance on oral antiplatelet agents to reduce complications of in-stent restenosis/thrombosis. Clopidogrel was initially introduced in 1997, and within the past seven years, two additional oral antiplatelet agents have been approved by the U.S. Food and Drug Administration. With more potent antiplatelet agents comes increased risks of adverse effects...
October 2016: South Dakota Medicine: the Journal of the South Dakota State Medical Association
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