Document Detail

Global amplification of sense RNA: a novel method to replicate and archive mRNA for gene expression analysis.
MedLine Citation:
PMID:  13679029     Owner:  NLM     Status:  MEDLINE    
We have developed a procedure to amplify mRNA into sense RNA (sRNA) so as to create a regenerating biorepository representing the complex mRNA profile in the original sample. The procedure exploits the template-switching activity of reverse transcriptase to incorporate RNA polymerase binding sites upstream of single-stranded cDNA (ss cDNA). Limited PCR was used for double-stranded DNA (dsDNA) synthesis. sRNA was synthesized from PCR products by in vitro transcription (IVT). sRNA was evaluated by real-time reverse transcription (RT)-PCR. sRNA synthesis was successful with RNA from human cell lines and tissues, yielding 2000- to 2500-fold amplification of glyceraldeyde-3 phosphate dehydrogenase (G3PDH). The size of sRNA ranged from 3.0 to 0.1 kb. sRNA synthesis preserved the relative differences in plant mRNAs spiked at abundance ranging over 5 orders of magnitude (0.00001-0.1%). This reflects the high fidelity of sRNA synthesis for mRNA as low as 0.3 copies/cell. sRNA is amplified synthetic mRNA in the 5'-->3' direction; the appropriate template for any gene expression analysis.
Mangalathu S Rajeevan; Irina M Dimulescu; Suzanne D Vernon; Mukesh Verma; Elizabeth R Unger
Related Documents :
24854859 - Vimentin knockdown decreases corneal opacity.
7687449 - Internal crna standards for quantitative northern analysis.
8068339 - Kinetics of hybridization of mrna of c-myc oncogene with 111in-labeled antisense oligod...
19747379 - Genome-wide expression profiling of in vivo-derived bloodstream parasite stages and dyn...
22923889 - Altered expression of hsp70 in oral lichen planus.
9054129 - Detection of feline immunodeficiency virus transcripts by quantitative reverse transcri...
19959649 - Expression, localization, and functional coupling of the somatostatin receptor subtype ...
20040059 - Delayed gastric emptying and disruption of the interstitial cells of cajal network afte...
17121089 - Cyclooxygenase-2 expression in feline pancreatic adenocarcinomas.
Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Genomics     Volume:  82     ISSN:  0888-7543     ISO Abbreviation:  Genomics     Publication Date:  2003 Oct 
Date Detail:
Created Date:  2003-09-18     Completed Date:  2004-06-01     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  8800135     Medline TA:  Genomics     Country:  United States    
Other Details:
Languages:  eng     Pagination:  491-7     Citation Subset:  IM    
Viral Exanthems and Herpesvirus Branch, Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, US Department of Health and Human Services, Atlanta, GA 30333, USA.
Export Citation:
APA/MLA Format     Download EndNote     Download BibTex
MeSH Terms
Cell Line
DNA, Complementary / biosynthesis
Gene Amplification
Gene Expression
Gene Expression Profiling / methods*
RNA / biosynthesis*
RNA, Messenger*
RNA-Directed DNA Polymerase
Reverse Transcriptase Polymerase Chain Reaction
Grant Support
Y1-CN-0101-01/CN/NCI NIH HHS
Reg. No./Substance:
0/DNA, Complementary; 0/RNA, Messenger; 63231-63-0/RNA; EC DNA Polymerase

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

Previous Document:  Cloning, characterization, and mRNA expression analysis of novel human fetal cochlear cDNAs.
Next Document:  STI-571: an anticancer protein-tyrosine kinase inhibitor.