Identification of differentially expressed genes in normal mucosa, adenomaand adenocarcinoma of colon by SSH

AIM To construct subtracted cDNA libraries and further identify differentia lly expressed genes that are related to the development of colorectal carci noma (CRC). METHODS Suppression subtractive hybridization (SSH) was done on cDNAs of no rmal mucosa, adenoma and adenocarcinoma tissues from the same patient. Thre e subtracted cDNA libraries were constructed and then hybridized with forwa rd and backward subtracted probes for differential screening. Positive clon es from each subtracted cDNA library were selected for sequencing and BLAST analysis. Finally, virtual Northern Blot confirmed such differential expre ssion. RESULTS By this way, there were about 3 - 4 x 10(2) clones identified in ea ch subtracted cDNA library, in which about 85% positive clones were differe ntially screened. Sequencing and BLAST homology search revealed some clones containing sequences of known gene fragments and several possibly novel ge nes showing few or no sequence homologies with any known sequences in the d atabase. CONCLUSION All results confirmed the effectiveness and sensitivity of SSH. The differentially expressed genes during the development of CRC can be use d to shed light on the pathogenesis of CRC and be useful genetic markers fo r early diagnosis and therapy.