SEQanswers

Go Back   SEQanswers > Literature Watch



Similar Threads
Thread Thread Starter Forum Replies Last Post
PubMed: RNA-ligase-dependent biases in miRNA representation in deep-sequenced small R Newsbot! Literature Watch 0 10-18-2011 02:00 AM
PubMed: Amplification Methods Bias Metagenomic Libraries of Uncultured Single-Strande Newsbot! Literature Watch 0 09-20-2011 02:00 AM
PubMed: Barcoded primers used in multiplex amplicon pyrosequencing bias amplification Newsbot! Literature Watch 0 09-06-2011 02:00 AM
PubMed: Bias Detection and Correction in RNA-Sequencing Data. Newsbot! Literature Watch 0 07-21-2011 02:00 AM
Hi, everybody, Do you agree amplification will delivery bias to RNA-seq seattle Introductions 3 05-19-2010 03:35 AM

Reply
 
Thread Tools
Old 01-11-2012, 02:30 AM   #1
Newsbot!
RSS Posting Maniac
 

Join Date: Feb 2008
Posts: 1,443
Default PubMed: Digital RNA sequencing minimizes sequence-dependent bias and amplification no

Syndicated from PubMed RSS Feeds

Digital RNA sequencing minimizes sequence-dependent bias and amplification noise with optimized single-molecule barcodes.

Proc Natl Acad Sci U S A. 2012 Jan 9;

Authors: Shiroguchi K, Jia TZ, Sims PA, Xie XS

Abstract
RNA sequencing (RNA-Seq) is a powerful tool for transcriptome profiling, but is hampered by sequence-dependent bias and inaccuracy at low copy numbers intrinsic to exponential PCR amplification. We developed a simple strategy for mitigating these complications, allowing truly digital RNA-Seq. Following reverse transcription, a large set of barcode sequences is added in excess, and nearly every cDNA molecule is uniquely labeled by random attachment of barcode sequences to both ends. After PCR, we applied paired-end deep sequencing to read the two barcodes and cDNA sequences. Rather than counting the number of reads, RNA abundance is measured based on the number of unique barcode sequences observed for a given cDNA sequence. We optimized the barcodes to be unambiguously identifiable, even in the presence of multiple sequencing errors. This method allows counting with single-copy resolution despite sequence-dependent bias and PCR-amplification noise, and is analogous to digital PCR but amendable to quantifying a whole transcriptome. We demonstrated transcriptome profiling of Escherichia coli with more accurate and reproducible quantification than conventional RNA-Seq.


PMID: 22232676 [PubMed - as supplied by publisher]



More...
Newsbot! is offline   Reply With Quote
Reply

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off




All times are GMT -8. The time now is 08:40 PM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2020, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO