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[Construction and screening of suppression subtractive hybridization library of renal cell carcinoma].

Yong Zhang, Junkui Ai, Zhiwen Zhang, Xiongjun Ye, Hong-jian Zhu, Dianqi Xin, Lili Liang, Yan-qun Na, Ying-lu Guo

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Abstract

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), 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. 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 its construction is the basis for further screening differentially expressed genes of RCC.

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What this paper is about

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), 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. 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 its construction is the basis for further screening differentially expressed genes of RCC.

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Available abstract

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), 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. 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 its construction is the basis for further screening differentially expressed genes of RCC.

Key concepts: Suppression subtractive hybridization, Gene, cDNA library, Complementary DNA, Molecular biology, Renal cell carcinoma, Genomic library, Biology

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[Construction and screening of suppression subtractive hybridization library of renal cell carcinoma]. — Research Paper | ScholarLens