Construction of a normalized cDNA library by introduction of a semi-solid mRNA-cDNA hybridization system.
We report a novel procedure to construct a normalized (equalized) cDNA library. By introduction of the highly efficient self-hybridization system between a whole mRNA population and their corresponding cDNA immobilized on latex beads, which involves relatively simple manipulations, we were able to generate an mRNA population in which the copy number of abundant species was reduced while that of rare species was enriched. In a typical experiment, after several cycles of self-hybridization on the beads, the ratio of the most to the least abundant marker mRNA species dropped by a factor of 300 (from 10,000 to 30) while the complexity and length of mRNAs in the population remained unchanged. The procedure should provide a potent tool for the expression cloning of cDNA and also facilitate the construction of whole cDNA catalogs from specific tissues (or cell types) from higher organisms.
Sasaki
Y F
YF
Institute of Molecular and Cellular Biosciences, University of Tokyo, Japan.
Ayusawa
D
D
Oishi
M
M
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D004351:Drug Resistance / Q000235:genetics; D015723:Gene Library; D005914:Globins / Q000235:genetics; D006801:Humans; D007700:Kinetics; D008863:Microspheres; D008969:Molecular Sequence Data; D009355:Neomycin; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics*; D011817:Rabbits
D018076:DNA, Complementary; D012333:RNA, Messenger; D005914:Globins; D009355:Neomycin