Objective To construct and screen the suppression subtractive hybridization (SSH) library of human renal cell carcinoma (RCC). Methods Poly A+ RNA was isolated from RCC lines 786-O(tester) and renal cell(RC) lines HK-...Objective To construct and screen the suppression subtractive hybridization (SSH) library of human renal cell carcinoma (RCC). Methods Poly A+ RNA was isolated from RCC lines 786-O(tester) and renal cell(RC) lines HK-2 ( driver), respectiely. SSH procedure was performed according to the protocol of the PCR-Select cDNA Subtraction Kit ( Clontech), and PCR products were cloned into pT-Adv vector and transformed E. coli TOP10F’. All positive clones picked out were digested and some of which were sequenced. Results The SSH library contained 362 clones with SSH cDNA fragments distributed mainly from 0.3 to 0.9 kb. Among 50 clones sequenced randomly,2 represented unknown genes and the other 48 derived from 36 known genes. Conclusion The quality of the SSH library of human RCC is reliable and is construction is the basis for further screening differentially expressed genes of RCC. 6 refs,4 figs, 1 tab.展开更多
文摘Objective To construct and screen the suppression subtractive hybridization (SSH) library of human renal cell carcinoma (RCC). Methods Poly A+ RNA was isolated from RCC lines 786-O(tester) and renal cell(RC) lines HK-2 ( driver), respectiely. SSH procedure was performed according to the protocol of the PCR-Select cDNA Subtraction Kit ( Clontech), and PCR products were cloned into pT-Adv vector and transformed E. coli TOP10F’. All positive clones picked out were digested and some of which were sequenced. Results The SSH library contained 362 clones with SSH cDNA fragments distributed mainly from 0.3 to 0.9 kb. Among 50 clones sequenced randomly,2 represented unknown genes and the other 48 derived from 36 known genes. Conclusion The quality of the SSH library of human RCC is reliable and is construction is the basis for further screening differentially expressed genes of RCC. 6 refs,4 figs, 1 tab.
