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学科主题: 临床医学
题名:
Suppression subtractive hybridization for identifying differentially expressed genes in renal cell carcinoma
作者: Zhang, Q; Zhang, ZW; Xin, DQ; Na, YQ; Guo, YL
关键词: kidney neoplasms ; carcinoma ; suppression subtractive hybridization ; library ; gene ; clone
刊名: CHINESE MEDICAL JOURNAL
发表日期: 2001-08-01
卷: 114, 期:8, 页:807-812
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology
类目[WOS]: Medicine, General & Internal
研究领域[WOS]: General & Internal Medicine
关键词[WOS]: ESTROGEN-RECEPTOR ; C-MYC ; CANCER
英文摘要:

Objective To construct a renal cell carcinoma ( RCC) cDNA subtractive library using suppression subtractive hybridization.

Methods Polyadenylated RNA [Poly (A)(+) RNA] was isolated from tissues of RCC and normal kidney, and single-strand cDNAs and double-strand cDNAs were synthesized in turn. RCC cDNAs were divided into two groups and ligated to the specific adaptors 1 and 2, and then hybridized with normal kidney cDNA twice with two rounds of suppression PCR. Second round PCR products were cloned to T/A plasmid vectors to set up the subtractive library. One hundred clones were randomly picked to perform enzyme digest analysis, and some underwent sequence analysis and Northern blot to identify RCC specifically expressed genes. SMART RACE procedure was operated to clone full length novel RCC specifically expressed genes.

Results A human RCC subtractive library with high subtractive efficiency was successfully set up. The amplified library contains 350 positive clones. Random analysis of 100 clones with enzyme restriction showed that 85 plasmids in the clones contained 50-400 bp inserts. Sequence analysis was performed for 10 clones. All the 10 sequences were unknown before and derived from 6 unique, novel genes among which the cDNA insert RCC18 had five copies. Northern blot analysis showed that RCC18 cDNA was highly expressed in RCC, but no signal could be detected in normal kidney. Using SMART RACE technique, we obtained the full length of the novel gene RCC18.

Conclusions The constructed cDNA subtractive library of human RCC is a highly efficient one and lays a solid foundation for large scale screening and cloning new and specific oncogenes or tumor suppressor genes of RCC. The novel specifically expressed genes provided an important clue for studying the mechanisms of occurrence and development of RCC.

语种: 英语
WOS记录号: WOS:000170838400006
Citation statistics:
内容类型: 期刊论文
URI标识: http://ir.bjmu.edu.cn/handle/400002259/63042
Appears in Collections:北京大学第一临床医学院_泌尿外科_期刊论文

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作者单位: 1.Beijing Med Univ, Affiliated Hosp 1, Inst Urol, Dept Urol, Beijing 100034, Peoples R China
2.Beijing Med Univ, Dept Physiol, Beijing 100083, Peoples R China

Recommended Citation:
Zhang, Q,Zhang, ZW,Xin, DQ,et al. Suppression subtractive hybridization for identifying differentially expressed genes in renal cell carcinoma[J]. CHINESE MEDICAL JOURNAL,2001,114(8):807-812.
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