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Sequence library construction

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https://www.readbyqxmd.com/read/28818039/the-impact-of-rna-sequence-library-construction-protocols-on-transcriptomic-profiling-of-leukemia
#1
Ashwini Kumar, Matti Kankainen, Alun Parsons, Olli Kallioniemi, Pirkko Mattila, Caroline A Heckman
BACKGROUND: RNA sequencing (RNA-seq) has become an indispensable tool to identify disease associated transcriptional profiles and determine the molecular underpinnings of diseases. However, the broad adaptation of the methodology into the clinic is still hampered by inconsistent results from different RNA-seq protocols and involves further evaluation of its analytical reliability using patient samples. Here, we applied two commonly used RNA-seq library preparation protocols to samples from acute leukemia patients to understand how poly-A-tailed mRNA selection (PA) and ribo-depletion (RD) based RNA-seq library preparation protocols affect gene fusion detection, variant calling, and gene expression profiling...
August 17, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28815506/preparing-fosmid-mate-paired-libraries-using-cre-loxp-recombination
#2
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/28813447/the-use-of-high-throughput-small-rna-sequencing-reveals-differentially-expressed-micrornas-in-response-to-aster-yellows-phytoplasma-infection-in-vitis-vinifera-cv-chardonnay
#3
Marius C Snyman, Marie-Chrystine Solofoharivelo, Rose Souza-Richards, Dirk Stephan, Shane Murray, Johan T Burger
Phytoplasmas are cell wall-less plant pathogenic bacteria responsible for major crop losses throughout the world. In grapevine they cause grapevine yellows, a detrimental disease associated with a variety of symptoms. The high economic impact of this disease has sparked considerable interest among researchers to understand molecular mechanisms related to pathogenesis. Increasing evidence exist that a class of small non-coding endogenous RNAs, known as microRNAs (miRNAs), play an important role in post-transcriptional gene regulation during plant development and responses to biotic and abiotic stresses...
2017: PloS One
https://www.readbyqxmd.com/read/28807055/transcriptomic-profiling-of-microbe-microbe-interactions-reveals-the-specific-response-of-the-biocontrol-strain-p-fluorescens-in5-to-the-phytopathogen-rhizoctonia-solani
#4
Rosanna C Hennessy, Mikkel A Glaring, Stefan Olsson, Peter Stougaard
BACKGROUND: Few studies to date report the transcriptional response of biocontrol bacteria toward phytopathogens. In order to gain insights into the potential mechanism underlying the antagonism of the antimicrobial producing strain P. fluorescens In5 against the phytopathogens Rhizoctonia solani and Pythium aphanidermatum, global RNA sequencing was performed. METHODS: Differential gene expression profiling of P. fluorescens In5 in response to either R. solani or P...
August 10, 2017: BMC Research Notes
https://www.readbyqxmd.com/read/28806924/essential-genome-of-campylobacter-jejuni
#5
Rabindra K Mandal, Tieshan Jiang, Young Min Kwon
BACKGROUND: Campylobacter species are a leading cause of bacterial foodborne illness worldwide. Despite the global efforts to curb them, Campylobacter infections have increased continuously in both developed and developing countries. The development of effective strategies to control the infection by this pathogen is warranted. The essential genes of bacteria are the most prominent targets for this purpose. In this study, we used transposon sequencing (Tn-seq) of a genome-saturating library of Tn5 insertion mutants to define the essential genome of C...
August 14, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28804478/the-entomopathogenic-fungi-isaria-fumosorosea-plays-a-vital-role-in-suppressing-the-immune-system-of-plutella-xylostella-rna-seq-and-dge-analysis-of-immunity-related-genes
#6
Jin Xu, Xiaoxia Xu, Muhammad Shakeel, Shuzhong Li, Shuang Wang, Xianqiang Zhou, Jialin Yu, Xiaojing Xu, Xiaoqiang Yu, Fengliang Jin
Most, if not all, entomopathogenic fungi have been used as alternative control agents to decrease the insect resistance and harmful effects of the insecticides on the environment. Among them, Isaria fumosorosea has also shown great potential to control different insect pests. In the present study, we explored the immune response of P. xylostella to the infection of I. fumosorosea at different time points by using RNA-Sequencing and differential gene expression technology at the genomic level. To gain insight into the host-pathogen interaction at the genomic level, five libraries of P...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/28801904/constructing-strong-cell-type-specific-promoters-through-informed-design
#7
Adam J Brown, David C James
Promoter functionality is highly context dependent, as exemplified by gene-specific expression profiles across different tissues and cell types. Cell type-specific promoter regulation is a function of each cell's unique complement of transcriptional machinery components. Accordingly, to achieve high levels of transcriptional activity within a particular cell type, synthetic promoters must be specifically designed to harness those cells discrete repertoire of available transcription factors . Here, we describe a method for constructing very strong cell type-specific synthetic promoters for use in any given mammalian host cell...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28801902/bioinformatically-informed-design-of-synthetic-mammalian-promoters
#8
Michael L Roberts, Polyxeni Katsoupi, Vivian Tseveleki, Era Taoufik
Synthetic promoters have been developed in a number of different organisms and are capable of mediating specific and enhanced levels of gene expression. Typically, cis-regulatory regions from a few genes are randomly combined to generate a synthetic promoter library, and the sequences with the highest activity are selected for in target cell lines. Here we describe a novel approach that can be employed in the construction of synthetic promoters . Specifically, we use gene expression profiles obtained from microarray datasets to select the cis-regulatory elements that comprise the synthetic promoter library...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28800965/the-crispr-cas9-facilitated-multiplex-pathway-optimization-cfpo-technique-and-its-application-to-improve-the-escherichia-coli-xylose-utilization-pathway
#9
Xinna Zhu, Dongdong Zhao, Huanna Qiu, Feiyu Fan, Shuli Man, Changhao Bi, Xueli Zhang
One of the most important research subjects of metabolic engineering is the pursuit of balanced metabolic pathways, which requires the modulation of expression of many genes. However, simultaneously modulating multiple genes on the chromosome remains challenging in prokaryotic organisms, including the industrial workhorse - Escherichia coli. In this work, the CRISPR/Cas9-facilitated multiplex pathway optimization (CFPO) technique was developed to simultaneously modulate the expression of multiple genes on the chromosome...
August 9, 2017: Metabolic Engineering
https://www.readbyqxmd.com/read/28791211/development-and-characterization-of-polymorphic-est-based-ssr-markers-in-barley-hordeum-vulgare
#10
Won-Sam Jo, Hye-Yeong Kim, Kyung-Min Kim
In barley, breeding using good genetic characteristics can improve the quality or quantity of crop characters from one generation to the next generation. The development of effective molecular markers in barley is crucial for understanding and analyzing the diversity of useful alleles. In this study, we conducted genetic relationship analysis using expressed sequence tag-simple sequence repeat (EST-SSR) markers for barley identification and assessment of barley cultivar similarity. Seeds from 82 cultivars, including 31 each of naked and hulled barley from the Korea Seed and Variety Service and 20 of malting barley from the RDA-Genebank Information Center, were analyzed in this study...
August 2017: 3 Biotech
https://www.readbyqxmd.com/read/28790814/bacteriological-incidence-in-pneumonia-patients-with-pulmonary-emphysema-a-bacterial-floral-analysis-using-the-16s-ribosomal-rna-gene-in-bronchoalveolar-lavage-fluid
#11
Keisuke Naito, Kei Yamasaki, Kazuhiro Yatera, Kentaro Akata, Shingo Noguchi, Toshinori Kawanami, Kazumasa Fukuda, Takashi Kido, Hiroshi Ishimoto, Hiroshi Mukae
Pulmonary emphysema is an important radiological finding in chronic obstructive pulmonary disease patients, but bacteriological differences in pneumonia patients according to the severity of emphysematous changes have not been reported. Therefore, we evaluated the bacteriological incidence in the bronchoalveolar lavage fluid (BALF) of pneumonia patients using cultivation and a culture-independent molecular method. Japanese patients with community-acquired pneumonia (83) and healthcare-associated pneumonia (94) between April 2010 and February 2014 were evaluated...
2017: International Journal of Chronic Obstructive Pulmonary Disease
https://www.readbyqxmd.com/read/28789614/comparative-transcriptome-analysis-reveals-key-cadmium-transport-related-genes-in-roots-of-two-pak-choi-brassica-rapa-l-ssp-chinensis-cultivars
#12
Rugang Yu, Dan Li, Xueling Du, Shenglan Xia, Caifeng Liu, Gangrong Shi
BACKGROUND: Cadmium translocation from roots to shoots is a complex biological process that is controlled by gene regulatory networks. Pak choi exhibits wide cultivar variations in Cd accumulation. However, the molecular mechanism involved in cadmium translocation and accumulation is still unclear. To isolate differentially expressed genes (DEGs) involved in transporter-mediated regulatory mechanisms of Cd translocation in two contrasting pak choi cultivars, Baiyewuyueman (B, high Cd accumulator) and Kuishan'aijiaoheiye (K, low Cd accumulator), eight cDNA libraries from the roots of two cultivars were constructed and sequenced by RNA-sequencing...
August 8, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28771149/culture-independent-methods-reveal-high-diversity-of-oxa-48-like-genes-in-water-environments
#13
Marta Tacão, Isabel Silva, Isabel Henriques
The carbapenemase OXA-48 was identified for the first time in 2001 and is now one of the greatest concerns in terms of antibiotic resistance. While many studies report clinical OXA-48-like producers, few reports refer blaOXA-48-like genes in environmental bacteria. The main goal of this study was to evaluate the diversity of blaOXA-48-like genes in aquatic systems, using culture-independent approaches. For that, environmental DNA was obtained from riverine and estuarine water and used to construct clone libraries of blaOXA-48-like gene polymerase chain reaction amplicons...
August 2017: Journal of Water and Health
https://www.readbyqxmd.com/read/28768814/construction-and-optimization-of-large-gene-co-expression-network-in-maize-using-rna-seq-data
#14
Ji Huang, Stefania Vendramin Alegre, Lizhen Shi, Karen McGinnis
With the emergence of massively parallel sequencing, genome-wide expression data production has reached an unprecedented level. This abundance of data has greatly facilitated maize research, but may not be amenable to traditional analysis techniques that were optimized for other data types. Using publicly available data, a Gene Co-expression Network (GCN) can be constructed and used for gene function prediction, candidate gene selection and improving understanding of regulatory pathways. Several GCN studies have been done in maize, mostly using microarray datasets...
August 2, 2017: Plant Physiology
https://www.readbyqxmd.com/read/28760689/comprehensive-discovery-of-non-coding-rnas-in-acute-myeloid-leukemia-cell-transcriptomes
#15
Jin Zhang, Malachi Griffith, Christopher A Miller, Obi L Griffith, David H Spencer, Jason R Walker, Vincent Magrini, Sean D McGrath, Amy Ly, Nichole M Helton, Maria Trissal, Daniel C Link, Ha X Dang, David E Larson, Shashikant Kulkarni, Matthew G Cordes, Catrina C Fronick, Robert S Fulton, Jeffery M Klco, Elaine R Mardis, Timothy J Ley, Richard K Wilson, Christopher A Maher
To comprehensively detect diverse and novel RNA species, we compared deep small RNA and RNA sequencing (RNA-seq) methods applied to a primary acute myeloid leukemia (AML) sample. We were able to discover previously unannotated small RNAs using deep sequencing of a library method employing broader insert size selection. We analyzed the long non-coding RNA (lncRNA) landscape in AML by comparing deep sequencing from multiple RNA-seq library construction methods for the sample we studied, then integrating RNA-seq data from 179 AML cases...
July 28, 2017: Experimental Hematology
https://www.readbyqxmd.com/read/28759014/functional-annotation-of-chemical-libraries-across-diverse-biological-processes
#16
Jeff S Piotrowski, Sheena C Li, Raamesh Deshpande, Scott W Simpkins, Justin Nelson, Yoko Yashiroda, Jacqueline M Barber, Hamid Safizadeh, Erin Wilson, Hiroki Okada, Abraham A Gebre, Karen Kubo, Nikko P Torres, Marissa A LeBlanc, Kerry Andrusiak, Reika Okamoto, Mami Yoshimura, Eva DeRango-Adem, Jolanda van Leeuwen, Katsuhiko Shirahige, Anastasia Baryshnikova, Grant W Brown, Hiroyuki Hirano, Michael Costanzo, Brenda Andrews, Yoshikazu Ohya, Hiroyuki Osada, Minoru Yoshida, Chad L Myers, Charles Boone
Chemical-genetic approaches offer the potential for unbiased functional annotation of chemical libraries. Mutations can alter the response of cells in the presence of a compound, revealing chemical-genetic interactions that can elucidate a compound's mode of action. We developed a highly parallel, unbiased yeast chemical-genetic screening system involving three key components. First, in a drug-sensitive genetic background, we constructed an optimized diagnostic mutant collection that is predictive for all major yeast biological processes...
September 2017: Nature Chemical Biology
https://www.readbyqxmd.com/read/28756139/a-universal-phage-display-system-for-the-seamless-construction-of-fab-libraries
#17
Renae S Nelson, Philippe Valadon
The construction of Fab phage libraries requires the cloning of domains from both the light and the heavy chain of antibodies. Despite the advent of powerful strategies such as splicing-by-overlap extension PCR, obtaining high quality libraries with excellent coverage remains challenging. Here, we explored the use of type IIS restriction enzymes for the seamless cloning of Fab libraries. We analyzed human, murine and rabbit germline antibody repertoires and identified combinations of restriction enzymes that exhibit very few or no recognition sites in the antibody sequences...
July 26, 2017: Journal of Immunological Methods
https://www.readbyqxmd.com/read/28755696/identification-and-functional-analysis-of-a-new-glyphosate-resistance-gene-from-a-fungus-cdna-library
#18
Bo Tao, Bai-Hui Shao, Yu-Xin Qiao, Xiao-Qin Wang, Shu-Jun Chang, Li-Juan Qiu
Glyphosate is a widely used broad spectrum herbicide; however, this limits its use once crops are planted. If glyphosate-resistant crops are grown, glyphosate can be used for weed control in crops. While several glyphosate resistance genes are used in commercial glyphosate tolerant crops, there is interest in identifying additional genes for glyphosate tolerance. This research constructed a high-quality cDNA library form the glyphosate-resistant fungus Aspergillus oryzae RIB40 to identify genes that may confer resistance to glyphosate...
August 2017: Pesticide Biochemistry and Physiology
https://www.readbyqxmd.com/read/28754632/small-rna-profiling-reveals-important-roles-for-mirnas-in-arabidopsis-response-to-bacillus-velezensis-fzb42
#19
Shanshan Xie, Haiyang Jiang, Zhilan Xu, Qianqian Xu, Beijiu Cheng
Bacillus velezensis FZB42 (previously classified as Bacillus amyloliquefaciens FZB42) has been confirmed to successfully colonize plant roots and enhance defense response against pathogen infection. This study indicated that FZB42 inoculation enhanced Arabidopsis defense response against Pseudomonas syringae DC3000 through inducing the expression of PR1, PDF1.2 and stomata closure. To further clarify the induced defense response at miRNA level, sRNA libraries from Arabidopsis roots inoculated with FZB42 and control were constructed and sequenced...
July 25, 2017: Gene
https://www.readbyqxmd.com/read/28754135/retrofitting-the-bac-cloning-vector-pbelobac11-by-the-insertion-of-a-mutant-loxp-site
#20
Jonathon S Coren
OBJECTIVE: Human genomic libraries constructed in bacterial artificial chromosome vectors were utilized to make physical maps of all 23-chromosome pairs and as the templates for DNA sequencing to aid in the completion of the Human Genome Project. The goal of this study was to modify the BAC vector pBeloBAC11 so that genomic inserts contained in this vector could be subjected to bidirectional transposon-mediated nested deletions from the wild-type and mutant loxP sites present. RESULTS: An oligonucleotide containing a mutant loxP 2272 site and a XhoI restriction enzyme sequence was designed and inserted at the SfiI restriction site located approximately 200 basepairs upstream of the lacZ gene in pBeloBAC11...
July 28, 2017: BMC Research Notes
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