SEQanswers

Go Back   SEQanswers > Literature Watch



Similar Threads
Thread Thread Starter Forum Replies Last Post
PubMed: Target-Enrichment Through Amplification of Hairpin-Ligated Universal Targets Newsbot! Literature Watch 0 03-25-2011 06:10 AM
PubMed: Characterization of transcriptional complexity during berry development in Vi Newsbot! Literature Watch 0 02-02-2010 02:10 AM
PubMed: Outcome prediction and risk assessment by quantitative pyrosequencing methyla Newsbot! Literature Watch 0 07-25-2009 05:07 AM
PubMed: Quantitative analysis of DNA methylation after whole bisulfitome amplificatio Newsbot! Literature Watch 0 05-22-2009 05:00 AM
PubMed: The development and impact of 454 sequencing. Newsbot! Literature Watch 0 10-11-2008 05:09 AM

Reply
 
Thread Tools
Old 03-12-2010, 02:00 AM   #1
Newsbot!
RSS Posting Maniac
 

Join Date: Feb 2008
Posts: 1,443
Default PubMed: Development and quantitative analyses of a universal rRNA-subtraction protoco

Syndicated from PubMed RSS Feeds

Related Articles Development and quantitative analyses of a universal rRNA-subtraction protocol for microbial metatranscriptomics.

ISME J. 2010 Mar 11;

Authors: Stewart FJ, Ottesen EA, Delong EF


Metatranscriptomes generated by pyrosequencing hold significant potential for describing functional processes in complex microbial communities. Meeting this potential requires protocols that maximize mRNA recovery by reducing the relative abundance of ribosomal RNA, as well as systematic comparisons to identify methodological artifacts and test for reproducibility across data sets. Here, we implement a protocol for subtractive hybridization of bacterial rRNA (16S and 23S) that uses sample-specific probes and is applicable across diverse environmental samples. To test this method, rRNA-subtracted and unsubtracted transcriptomes were sequenced (454 FLX technology) from bacterioplankton communities at two depths in the oligotrophic open ocean, yielding 10 data sets representing approximately 350 Mbp. Subtractive hybridization reduced bacterial rRNA transcript abundance by 40-58%, increasing recovery of non-rRNA sequences up to fourfold (from 12% to 20% of total sequences to 40-49%). In testing this method, we established criteria for detecting sequences replicated artificially via pyrosequencing errors and identified such replicates as a significant component (6-39%) of total pyrosequencing reads. Following replicate removal, statistical comparisons of reference genes (identified via BLASTX to NCBI-nr) between technical replicates and between rRNA-subtracted and unsubtracted samples showed low levels of differential transcript abundance (
Newsbot! is offline   Reply With Quote
Old 03-12-2010, 05:41 PM   #2
ECO
--Site Admin--
 
Location: SF Bay Area, CA, USA

Join Date: Oct 2007
Posts: 1,358
Default

Ah...my prior life.
ECO 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 10:12 PM.


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