恶性血液病细胞株人端粒重复序列结合因子基因突变情况的研究

目的　检测 1 1个端粒酶阳性的恶性血液病细胞株中人端粒重复序列结合因子 1(TERF1 )基因的突变情况并探讨其意义。方法　用生物医学软件预测TERF1基因组结构并用PCR和测序的方法证实 ,确定TERF1全长基因组结构。采用端粒重复序列扩增 (TRAP) 酶联免疫吸附实验 (ELISA)检测细胞株的端粒酶活性。以PCR产物直接测序的方法检测髓系白血病细胞株K5 6 2、HL 6 0、U937、NB4、THP 1、HEL、Dami,淋巴细胞白血病细胞株 6T CEM、Jurkat和Raji,以及骨髓增生异常综合征 (MDS RAEB)细胞株MUTZ 1中TERF1外显子的突变情况 ,以正常人骨髓单个核细胞作对照。结果　TERF1基因组全长 38.6kb ,由 1 0个外显子和 9个内含子构成 ,外显子和内含子的接合边界获确认。 1 1种恶性血液病细胞株均呈端粒酶阳性。在所检测的细胞株中未发现外显子存在有意义的突变 ,但在外显子周边的部分内含子中发现 4个位点有变异 ,其中MUTZ 1的变异有别于白血病细胞株 ,未发现髓系和淋巴细胞系细胞株之间的变异情况有差异。结论　TERF1外显子突变可能并不是恶性血液病细胞中TERF1功能异常的主要原因

Study on the mutation of human telomeric repeat binding factor 1 gene in malignant hematopoietic cell lines

Objective To detect mutations of human telomeric repeat binding factor 1 (TERF1) gene in 11 malignant hematopoietic cell lines, which have positive telomerase activity, and evaluate the significance of the mutations. Methods Genome structure of TERF1 was predicted by using biology information program, and verified by PCR and sequencing. Telomerase activity was detected by telomeric repeat amplification(TRAP) ELISA. PCR and sequencing were used to detect mutation of each exon of TERF1 in 11 cell lines, including myelogenous leukemia cell lines K562, HL 60, U 937, NB4, THP 1, HEL and Dami; lymphoblastic leukemia cell lines 6T CEM, Jurkat and Raji and MDS RAEB cell line MUTZ 1. Five DNA samples from healthy volunteers were detected as normal controls. Results TERF1 gene has 10 exons and spans 38.6 kb. All the 11 cell lines showed positive telomerase activity. No mutation was found in all exons of TERF1 in the 11 cell lines. However, 4 variants were found in intron1,2 and 8 near exon1,exon2 and exon9, respectively. The variants in MUTZ 1 was different from those in leukemia cell lines; but no difference was found between the variants in myelogenous and lymphoblastic leukemia cell lines. Conclusion TERF1 mutation is probably not among the main factors of the gene dysfunction in malignant hematopoietic diseases.