摘要
目的 筛选肺癌细胞恶性转化不同时期差异表达的基因,以期用于肺癌的诊断。方法 应用抑制消减杂交技术(SSH)、测序、cDNA芯片(cDNA Microarry)、northern blot。结果 利用SSH建立了永生化人支气管上皮细胞(BEP2D)恶性转化不同时期差异表达基因的cDNA文库,其中,A消减文库(BEP2D细胞的cDNA为tester,α粒子照射BEP2D细胞后35代恶性转化细胞R15Hp35的cDNA为driver)有416个克隆,B消减文库(α粒子照射BEP2D细胞后20代转化细胞R15Hp20的cDNA为tester,BEP2D和R15Hp35细胞的cDNA混合后为driver)有301个克隆,C消减文库(R15Hp35细胞的cDNA为tester,BEP2D细胞的cDNA为driver)有586个克隆。对文库中107个克隆单向测序发现:19个克隆在GenBank中没有查到对应的同源序列,其它88个克隆共代表了76个不同的已知基因。然后,将3个文库中全部克隆的cDNA制作成cDNA芯片,用该芯片筛选了15例肺癌组织、5例肺癌旁组织和其他8种癌组织(肝癌、胃癌、食管癌、乳腺癌、白血病、子宫内膜癌、脑神经角质瘤和结肠癌)中mRNA的表达差异。结果,获得肺癌组织高于肺癌旁组织表达的cDNA 26个,肺癌旁组织高于肺癌组织表达的31个,2者高于其他8种癌组织的分别为:肺癌旁组织中63个,肺癌组织中87个。将这208个具较大差异表达的基因重新制作成cDNA芯片,
Objective: Screen the differentially expressed genes in differentially malignant transformed lung cancer cell lines, in the hope of applying to lung cancer diagnosis. Methods: suppression subtractive hybridization (SSH), sequence, cDNA Microarray, northern blot. Result: Constructed three differentially expressed cDNA libraries from different malignant transformed human bronchial epithelial cells(BEP2D) using SSH: A. subtraction library (The cDNAs of BEP2D cells as tester and that of R15Hp35 cells as driver. ) contained 416 clones; B. subtraction library (The cDNAs of R15Hp20 cells as tester and that of R15Hp35 and BEP2D cells mixed together as driver. ) contained 301 clones; C. subtraction library (The cDNAs of R15Hp35 cells as tester and that of BEP2D cells as driver. ) contained 568 clones. Then, 107 cDNA clones were sequenced and analyzed: 19 cDNAs were found to be novel ones; and 88 cDNAs represent 76 different known genes. After that, the three library clones were made on one cDNA chip and hybridized with probes which come from 15 samples lung cancer tissues, 5 sample lung paracancerous tissues and other 8 kinds of tumor tissues (3 samples for each, including liver, gastric, esophageal, breast, leukaemia, endometrium, glioma, colorectal. ) respectively: 27 cDNAs expressed higher in lung cancer tissue than in paracancerous tissues; 31 cDNAs expressed higher in lung paracancerous tissues than in cancer tissues. Compared with other 8 kinds tumors, lung paracancerous tissues have 63 cDNAs and lung cancer tissues have 87 cDNAs over expressed. Put the 208 (27 +31 +63 + 87) cDNAs on one cDNA chip and hybridized with cancer and paracancerous tissues which come from same squamous cell carcinoma patient, the expression profile has distinct difference between the two tissues. Conclusion: The differentially expressed genes may be the potentially genes for diagnosis of lung cancer.
出处
《循证医学》
CSCD
2002年第2期75-79,共5页
The Journal of Evidence-Based Medicine
关键词
肺癌
诊断
基因
CDNA芯片
BEP2D细胞
lung cancer
diagnosis
genes
BEP2D cell
cDNA Microarray
suppression subtractive hybridization
