Suppression subtractive hybridization: A method for generating differentially regulated or tissue-specific cDNA probes and libraries

Luda Diatchenko, Yun Fai Chris Lau, Aaron P. Campbell, Alex Chenchik, Fauzia Moqadam, Betty Huang, Sergey Lukyanov, Konstantin Lukyanov, Nadya Gurskaya, Eugene D. Sverdlov, Paul D. Siebert

Research output: Contribution to journalArticlepeer-review

2725 Citations (Scopus)

Abstract

A new and highly effective method, termed suppression subtractive hybridization (SSH), has been developed for the generation of subtracted cDNA libraries. It is based primarily on a recently described technique called suppression PCR and combines normalization and subtraction in a single procedure. The normalization step equalizes the abundance of cDNAs within the target population and the subtraction step excludes the common sequences between the target and driver populations. In a model system, the SSH technique enriched fur rare sequences over 1,000-fold in one round of subtractive hybridization. We demonstrate its usefulness by generating a testis-specific cDNA library and by using the subtracted cDNA mixture as a hybridization probe to identify homologous sequences in a human Y chromosome cosmid library. The human DNA inserts in the isolated cosmids were further confirmed to be expressed in a testis-specific manner. These results suggest that the SSH technique is applicable to many molecular genetic and positional cloning studies for the identification of disease, developmental, tissue- specific, or other differentially expressed genes.

Original languageEnglish
Pages (from-to)6025-6030
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume93
Issue number12
DOIs
Publication statusPublished - 11 Jun 1996
Externally publishedYes

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