Semi-multiplex PCR technique for screening of abundant transcripts during systematic sequencing of cDNA libraries.
The systematic sequencing of cDNA libraries is an efficient approach for the identification of new genes, but the presence of abundant mRNAs is often a major problem. This paper describes a very simple method of "semi-multiplex PCR" that allows specific identification of such abundant transcripts before DNA sequencing without using nonrepresentative subtracted libraries. The PCR utilizes a series of forward primers specific for abundant transcripts with a pair of universal primers used for template generation. cDNA clones corresponding to abundant mRNAs are then revealed by double bands in agarose gel.
Pacchioni
B
B
CRIBI Biotechnology Centre, Università degli Studi di Padova, Italy.
Trevisan
S
S
Gomirato
S
S
Toppo
S
S
Valle
G
G
Lanfranchi
G
G
D001483:Base Sequence; D018076:DNA, Complementary / Q000737:chemistry*; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis*
D018076:DNA, Complementary; D012333:RNA, Messenger