新基因1A6定位及在肿瘤中的双色荧光原位杂交分析

Localization and analysis of 1A6 gene by dual color fluorescence in situ hybridization on gastric carcinoma tissue and tumor cell lines

目的 了解定位新基因 1A6在不同组织来源肿瘤中的状态。方法 用单色FISH法将载于质粒的 1A6基因小片段cDNA定位于C组某一染色体长臂 ,根据该基因测序结果设计 8对引物 ,通过PCR在PAC文库中筛选目的基因的基因组DNA。用切口翻译法分别用红绿荧光直接标记 12号α 卫星重复序列和 1A6基因组DNA ,采用双色荧光共杂交的方法定位 1A6基因 ,并对其及 12号染色体进行了数量分析。结果 1A6基因定位于 12q2 3 .2 2 3.3。 1A6基因在乳腺癌、卵巢癌及胃癌细胞株中双色FISH的绿红信号比 (R值 )为 0 .96～ 1.0 1,在胃癌组织印片中R值为 0 .93～ 1.11。 12号染色体在乳腺癌细胞株BMI中的双体率 87.7% ,三体率 7.4% ;在卵巢癌细胞株SKOV3中的多体率 10 0 % ,其中五体率 6 7.6 % ;在胃癌细胞株BGC82 3中的多体率为 83.1% ,其中三体率为 71%。在实体瘤胃癌组织印片中 ,86 .4% (19/ 2 2 )的病例 12号染色体以双拷贝为主。结论 1A6基因在实体瘤胃癌组织及肿瘤细胞株 (BGC82 3、SKOV3、BMI)中均无明显扩增和缺失 ,推测其在肿瘤中的作用方式不是扩增或缺失。 12号染色体数目在不同组织来源肿瘤细胞株中各异 ,意义有待探讨。

Objective To locate 1A6 gene on the chromosome and study its copy number by dual color fluorescence in situ hybridization (FISH) on three tumor cell lines and 22 gastric tumors patients.Methods The single color FISH of small size biotin labeled cDNA cloned in plasmid was used to locate the 1A6 gene on one of the chromosomes in C group. Genomic DNA of 1A6 gene was screened by polymerase chain reaction (PCR) in PAC library according to the sequence. The florensence of green and orange was incorporated into the target gene 1A6 cloned in PAC and chromosome 12 specific α satellite repeat DNA by nick translation, followed by dual color FISH. 1A6 gene was located in the metaphase chromosome of the normal peripheral lymphocytes. 1A6 gene and chromosome 12 copy numbers were analyzed on touch slide of gastric cancer tissue and cell lines.Results 1A6 gene was localated in 12q23.2 23.3. The chromosome 12/1A6 signal ratio was 0.96 1.01 in breast, ovarian and gastric cancer cell lines. The ratio was 0.93 1.11 in gastric cancer tissue touch slides. The number of chromosome 12 is disomic (87.7%),triploid (7.4%) in BMI cell line; multisomic(100%),including pentasomic (67.6%) in SKOV3;the multisomic (83.1%),including trisomic (71%) in SGC823.There are highly disomic rate in 86.4% (19/22)of patients.Conclusion 1A6 gene is in 12q23.2 23.3.Neither 1A6 amplification nor deletion in gastric cancer tissue and three cell lines was found. Further study is needed for the understanding of chromosome 12 copy number variation in different cancer cell lines. [