抑癌基因TSG101及其剪接变异体在正常人外周血细胞中的表达
发布时间：2019-04-29 17:40
【摘要】：目的:研究抑癌基因TSG101及其四种剪接变异体在健康女性外周血单个核细胞中的表达规律及突变,探讨该基因在乳腺癌发生发展过程中所扮演的角色。 方法:从健康女性外周血单个核细胞中提取总RNA,针对TSG101基因全长及其四种剪接变异体(V_A、V_B、V_C和V_D)设计5对特异引物进行RT-PCR,PCR产物进行琼脂糖凝胶电泳观察分析,TSG101基因全长和四种剪接变异体PCR产物进行测序;并从部分样本提取基因组DNA,运用TSG101基因的第4外显子特异引物,进行PCR,对其产物进行序列测定。 结果:1、在37例健康女性外周血中均扩增出了TSG101全长及其四种剪接变异体片段。其中V_A、V_B和V_C的表达较一致,全长和V_D的表达存在差别,部分样本全长表达明显减弱。2、测序结果显示37例中7例(18.92%)均在TSG101基因第4外显子(mRNA第798bp处)存在“C→T”杂合性点突变;另30例(81.08%)经序列比对与基因库的TSG101mRNA序列100%一致。后来在DNA水平上也证实了突变的存在。 结论:1、在健康女性外周血细胞中存在TSG101基因的全长及四种剪接变异体片段。2、TSG101基因在健康女性存在杂合性突变,国内外尚未系统报道。该突变对剪接变异体B产生了错义突变。3、TSG101基因的杂合性突变可能提示TSG101基因外显子内存在单核苷酸多态性。
[Abstract]:Aim: to study the expression and mutation of tumor suppressor gene TSG101 and its four splicing variants in peripheral blood mononuclear cells (PBMC) of healthy women, and to explore the role of the gene in the carcinogenesis and development of breast cancer. Methods: total RNA, was extracted from peripheral blood mononuclear cells of healthy women. Five pairs of specific primers were designed for the full length of TSG101 gene and its four splicing variants (V A, V B, V C and V D) for RT-PCR,. The PCR products were analyzed by agarose gel electrophoresis. The full length of TSG101 gene and the PCR products of four splicing variants were sequenced. Genomic DNA, was extracted from some samples and sequenced by PCR, using specific primers of exon 4 of TSG101 gene. Results: 1. The full length of TSG101 and its four splicing variants were amplified in the peripheral blood of 37 healthy women. Among them, the expression of V _ (A), V _ (B) and V _ (C) were the same, the expression of full length and V _ (?) D were different, and the expression of the whole length of some samples was obviously decreased. 2. The sequencing results showed that 7 of 37 cases (18.92%) had "C / T" heterozygosity point mutation in exon 4 of TSG101 gene (mRNA 798 BP). The other 30 cases (81.08%) were identical with the TSG101mRNA sequence of gene bank by sequence alignment. The mutation was later confirmed at the DNA level. Conclusion: 1. The full length of TSG101 gene and four splicing variants exist in the peripheral blood cells of healthy women. 2. The heterozygosity mutation of TSG101 gene in healthy women has not been reported systematically at home and abroad. This mutation produces missense mutation in splicing variant B. 3. The heterozygosity mutation of TSG101 gene may indicate the existence of single nucleotide polymorphism in exon of TSG101 gene.
【学位授予单位】：暨南大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R346
[Abstract]:Aim: to study the expression and mutation of tumor suppressor gene TSG101 and its four splicing variants in peripheral blood mononuclear cells (PBMC) of healthy women, and to explore the role of the gene in the carcinogenesis and development of breast cancer. Methods: total RNA, was extracted from peripheral blood mononuclear cells of healthy women. Five pairs of specific primers were designed for the full length of TSG101 gene and its four splicing variants (V A, V B, V C and V D) for RT-PCR,. The PCR products were analyzed by agarose gel electrophoresis. The full length of TSG101 gene and the PCR products of four splicing variants were sequenced. Genomic DNA, was extracted from some samples and sequenced by PCR, using specific primers of exon 4 of TSG101 gene. Results: 1. The full length of TSG101 and its four splicing variants were amplified in the peripheral blood of 37 healthy women. Among them, the expression of V _ (A), V _ (B) and V _ (C) were the same, the expression of full length and V _ (?) D were different, and the expression of the whole length of some samples was obviously decreased. 2. The sequencing results showed that 7 of 37 cases (18.92%) had "C / T" heterozygosity point mutation in exon 4 of TSG101 gene (mRNA 798 BP). The other 30 cases (81.08%) were identical with the TSG101mRNA sequence of gene bank by sequence alignment. The mutation was later confirmed at the DNA level. Conclusion: 1. The full length of TSG101 gene and four splicing variants exist in the peripheral blood cells of healthy women. 2. The heterozygosity mutation of TSG101 gene in healthy women has not been reported systematically at home and abroad. This mutation produces missense mutation in splicing variant B. 3. The heterozygosity mutation of TSG101 gene may indicate the existence of single nucleotide polymorphism in exon of TSG101 gene.
【学位授予单位】：暨南大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R346
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相关期刊论文 前6条
1 陈照立;P16抑癌基因研究进展及在口腔肿瘤中的作用[J];泸州医学院学报;2003年04期
2 郑莹,李德，
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