Display Settings:

Format

Send to:

Choose Destination
See comment in PubMed Commons below
BMC Genomics. 2004 Jun 15;5(1):36.

SMART amplification combined with cDNA size fractionation in order to obtain large full-length clones.

Author information

  • 1Department of Molecular Genome Analysis, German Cancer Research Center (DKFZ), Heidelberg, Germany. r.wellenreuther@dkfz.de

Abstract

BACKGROUND:

cDNA libraries are widely used to identify genes and splice variants, and as a physical resource for full-length clones. Conventionally-generated cDNA libraries contain a high percentage of 5'-truncated clones. Current library construction methods that enrich for full-length mRNA are laborious, and involve several enzymatic steps performed on mRNA, which renders them sensitive to RNA degradation. The SMART technique for full-length enrichment is robust but results in limited cDNA insert size of the library.

RESULTS:

We describe a method to construct SMART full-length enriched cDNA libraries with large insert sizes. Sub-libraries were generated from size-fractionated cDNA with an average insert size of up to seven kb. The percentage of full-length clones was calculated for different size ranges from BLAST results of over 12,000 5'ESTs.

CONCLUSIONS:

The presented technique is suitable to generate full-length enriched cDNA libraries with large average insert sizes in a straightforward and robust way. The representation of full-coding clones is high also for large cDNAs (70%, 4-10 kb), when high-quality starting mRNA is used.

PMID:
15198809
[PubMed - indexed for MEDLINE]
PMCID:
PMC436056
Free PMC Article

Images from this publication.See all images (5)Free text

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for BioMed Central Icon for PubMed Central
    Loading ...
    Write to the Help Desk