A Novel Gene-cloning Method-Subtractive Hybridization Difference Display

SHDD (subtractive hybridization difference display) is a newly developed gene-cloning method, which combines a highly specific subtractive hybridization of cDNA representational difference analysis (cDNA RDA) with a sensitive bi-directional radioactive detection of mRNA differential display (mRNA DD). The major characteristic of this modified method is adopting a first round of bi-directional subtractive hybridization between two sample cDNAs and an appropriate second round of bi-directional subtractive hybridization between the two resultant first-round difference products, followed by labeling resultant difference products with isotope and separating these difference products on DNA sequencing gel. All these modifications facilitate SHDD method in isolating mRNA species with very low expression levels and/or very small expression differences between two related sample cDNAs, and obtaining more difference products with increased specificity and work efficiency, compared with typical mRNA DD and cDNA RDA.