Construction and screening of suppression subtractive hybridization library of renal cell carcinoma.
- Author:
Yong ZHANG
1
;
Jun-kui AI
;
Zhi-wen ZHANG
;
Xiong-jun YE
;
Hong-jian ZHU
;
Dian-qi XIN
;
Li-li LIANG
;
Yan-qun NA
;
Ying-lu GUO
Author Information
- Publication Type:Journal Article
- MeSH: Adenocarcinoma, Clear Cell; genetics; Cell Line, Tumor; Gene Library; Humans; Kidney Neoplasms; genetics; Nucleic Acid Hybridization; methods
- From: Chinese Journal of Surgery 2003;41(2):90-92
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct and screen the suppression subtractive hybridization (SSH) library of human renal cell carcinoma (RCC).
METHODSPoly A(+) RNA was isolated from RCC lines 786-O (tester) and renal cell (RC) lines HK-2 (driver), respectively. 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.
RESULTSThe 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.
CONCLUSIONThe quality of the SSH library of human RCC is reliable and its construction is the basis for further screening differentially expressed genes of RCC.
