Prevalence and correlates of GB virus C infection in HIV-infected and HIV-uninfected pregnant women in Bangkok, Thailand.
AuthorsBhanich Supapol W, Remis RS, Raboud J, Millson M, Tappero J, Kaul R, Kulkarni P, McConnell MS, Mock PA, McNicholl JM, Roongpisuthipong A, Chotpitayasunondh T, Shaffer N, Butera S
JournalJ Med Virol
PubMed ID21108337
'GB virus C (GBV-C) is an apathogenic virus that has been shown to inhibit HIV replication. This study examined the prevalence and correlates of GBV-C infection and clearance in three cohorts of pregnant women in Thailand. The study population consisted of 1,719 (1,387 HIV-infected and 332 HIV-uninfected) women from three ... More
An in vitro DNA double-strand break repair assay based on end-joining of defined duplex oligonucleotides.
AuthorsDatta K, Purkayastha S, Neumann RD, Winters TA
JournalMethods Mol Biol
PubMed ID22941624
'DNA double-strand breaks (DSBs) are caused by endogenous cellular processes such as oxidative metabolism, or by exogenous events like exposure to ionizing radiation or other genotoxic agents. Repair of these DSBs is essential for the maintenance of cellular genomic integrity. In human cells, and cells of other higher eukaryotes, DSBs ... More
Smooth muscle phenotypic diversity is mediated through alterations in myocardin gene splicing.
AuthorsIlagan RM, Genheimer CW, Quinlan SF, Guthrie KI, Sangha N, Ramachandrannair S, Kelley RW, Presnell SC, Basu J, Ludlow JW
JournalJ Cell Physiol
PubMed ID21792927
'Myocardin (MYOCD) is a smooth and cardiac muscle-specific transcriptional coactivator that is required for the proper expression of contraction-related genes. Through its function to transactivate effector genes, MYOCD plays an essential role in mediating the switch between contractile and non-contractile phenotypes, particularly in smooth muscle cells (SMC). There are at ... More
Genome wide full-length transcript analysis using 5' and 3' paired-end-tag next generation sequencing (RNA-PET).
AuthorsRuan X, Ruan Y
JournalMethods Mol Biol
PubMed ID22113299
'RNA-PET is a paired end tag (PET) sequencing method for full-length mRNA transcripts analysis using the next generation sequencer platforms such as Illumina GA and SOLiD. Unlike RNA-Seq method that sequences randomly sheared shotgun RNA short fragments, RNA-PET captures and sequences the 5'' and 3'' end tags of full-length cDNA ... More
Rapid, low-input, low-bias construction of shotgun fragment libraries by high-density in vitro transposition.
AuthorsAdey A, Morrison HG, Asan Xun X, Kitzman JO, Turner EH, Stackhouse B, MacKenzie AP, Caruccio NC, Zhang X, Shendure J
JournalGenome Biol
PubMed ID21143862
We characterize and extend a highly efficient method for constructing shotgun fragment libraries in which transposase catalyzes in vitro DNA fragmentation and adaptor incorporation simultaneously. We apply this method to sequencing a human genome and find that coverage biases are comparable to those of conventional protocols. We also extend its ... More
Quantification of gene transcripts with deep sequencing analysis of gene expression (DSAGE) using 1 to 2 µg total RNA.
AuthorsChristodoulou DC, Gorham JM, Kawana M, DePalma SR, Herman DS, Wakimoto H
JournalCurr Protoc Mol Biol
PubMed ID21225638
Deep sequencing analysis of gene expression (DSAGE) measures global gene transcript levels from only 1 to 2 µg total RNA by massively parallel sequencing of cDNA tags. This unit describes the construction of 21-bp cDNA tag libraries appropriate for massively parallel sequencing and analysis of the resulting sequence data. The ... More
Regulation of miR-17-92a cluster processing by the microRNA binding protein SND1.
AuthorsHeinrich EM, Wagner J, Krüger M, John D, Uchida S, Weigand JE, Suess B, Dimmeler S
Journal
PubMed ID23770094
MicroRNAs are small non-coding RNAs that regulate gene expression. Although all seven members of the miR-17-92a cluster originate from one primary transcript they are differentially expressed suggesting the presence of posttranscriptional regulation. By RNA pulldown and mass spectrometry we identified SND1, a known regulator of edited RNAs, interacting with pre-miR-92a ... More
Advances in sequencing technology have led to the development of many high-resolution methodologies that observe genomic activity and gene expression. This unit describes such an approach, native elongating transcript sequencing (NET-seq), which reveals the density of RNA polymerase across the Saccharomyces cerevisiae genome with single-nucleotide resolution. A procedure for capturing ... More
Beta toxin catalyzes formation of nucleoprotein matrix in staphylococcal biofilms.
Biofilms are surface-associated communities of microbes encompassed by an extracellular matrix. It is estimated that 80% of all bacterial infections involve biofilm formation, but the structure and regulation of biofilms are incompletely understood. Extracellular DNA (eDNA) is a major structural component in many biofilms of the pathogenic bacterium Staphylococcus aureus, ... More
The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments.
Recent studies highlight the importance of translational control in determining protein abundance, underscoring the value of measuring gene expression at the level of translation. We present a protocol for genome-wide, quantitative analysis of in vivo translation by deep sequencing. This ribosome profiling approach maps the exact positions of ribosomes on ... More
Developmental expression of non-coding RNAs in Chlamydia trachomatis during normal and persistent growth.
AuthorsAbdelrahman YM, Rose LA, Belland RJ
JournalNucleic Acids Res
PubMed ID21051342
Chlamydia trachomatis is an obligate intracellular bacterium that exhibits a unique biphasic developmental cycle that can be disrupted by growth in the presence of IFN-? and ß-lactams, giving rise to an abnormal growth state termed persistence. Here we have examined the expression of a family of non-coding RNAs (ncRNAs) that ... More
Target-enrichment through amplification of hairpin-ligated universal targets for next-generation sequencing analysis.
AuthorsSingh P, Nayak R, Kwon YM
JournalMethods Mol Biol
PubMed ID21431777
With rapid development of next-generation sequencing (NGS) technologies, it is becoming increasingly feasible to sequence entire genomes of various organisms from virus to human. However, in many occasions, it is still more practical to sequence and analyze only small regions of the entire genome that are informative for the purpose ... More
RNA sequencing and quantitation using the Helicos Genetic Analysis System.
AuthorsRaz T, Causey M, Jones DR, Kieu A, Letovsky S, Lipson D, Thayer E, Thompson JF, Milos PM
JournalMethods Mol Biol
PubMed ID21431761
The recent transition in gene expression analysis technology to ultra high-throughput cDNA sequencing provides a means for higher quantitation sensitivity across a wider dynamic range than previously possible. Sensitivity of detection is mostly a function of the sheer number of sequence reads generated. Typically, RNA is converted to cDNA using ... More
The amino acid linker between the endonuclease and helicase domains of adeno-associated virus type 5 Rep plays a critical role in DNA-dependent oligomerization.
AuthorsMaggin JE, James JA, Chappie JS, Dyda F, Hickman AB
JournalJ Virol
PubMed ID22205752
The adeno-associated virus (AAV) genome encodes four Rep proteins, all of which contain an SF3 helicase domain. The larger Rep proteins, Rep78 and Rep68, are required for viral replication, whereas Rep40 and Rep52 are needed to package AAV genomes into preformed capsids; these smaller proteins are missing the site-specific DNA-binding ... More
Analysis of the polymerization initiation and activity of Pasteurella multocida heparosan synthase PmHS2, an enzyme with glycosyltransferase and UDP-sugar hydrolase activity.
AuthorsChavaroche AA, van den Broek LA, Springer J, Boeriu C, Eggink G
JournalJ Biol Chem
PubMed ID21084307
Heparosan synthase catalyzes the polymerization of heparosan (-4GlcUAß1-4GlcNAca1-)(n) by transferring alternatively the monosaccharide units from UDP-GlcUA and UDP-GlcNAc to an acceptor molecule. Details on the heparosan chain initiation by Pasteurella multocida heparosan synthase PmHS2 and its influence on the polymerization process have not been reported yet. By site-directed mutagenesis of ... More