中国人NF1基因突变分析

Study on mutations of NF1 gene in Chinese

目的通过对28个家系56例Ⅰ型神经纤维瘤病患者NF1-GRD相邻外显子进行突变检测,探讨和分析中国人NF1基因的突变规律、突变类型、机制以及与临床表型的相关性,以期发现潜在的突变热点。方法采用聚合酶链反应-单链构象多态性和(或)异源双链分析(SSCP/HA)技术,结合DNA测序方法,对56例Ⅰ型神经纤维瘤病患者的NF1基因第20、28、29、39号外显子的突变或多态性进行筛查。结果在所检测的28个家系中有一家系父子3例Ⅰ型神经纤维瘤病患者的NF1基因第20号外显子均出现异常移动的电泳条带,重复检测单链构象多态实验无异常发现,而异源双链实验则仍显示有1条移动异常的双链电泳条带,DNA测序证实为该家系第20号外显子上T→G的杂合性突变,即Leu1141Arg错义突变。同时发现,另1例Ⅰ型神经纤维瘤病患者的第28号外显子亦表现为电泳条带移动异常,测序证实为第28号外显子5'端上游第28位核苷酸G→T杂合子。未发现第29号和39号外显子电泳条带移动异常。结论聚合酶链反应-单链构象多态性与异源双链分析联合应用,可提高Ⅰ型神经纤维瘤病基因突变检测的敏感性和阳性检出率。所测28个家系56例患者NF1基因的第20、28、29及39号外显子并非突变热点或突变率相对较高的区域。

Objective Fifty-six neurofibromatosis type 1 （NF-1） eases from 28 families were screened for mutations in some exons of NF1 gene near NF1-GRD region. In this paper, we studied gene mutation reg- ularity, type, mechanism, and related clinical phenotype in Chinese NF-1 patients in order to find out the potential hotspots of NF1 gene mutation. Methods This study included 56 NF-1 patients. Mutation or polymorphism of exon 20, 28, 29, 39 in these patients were screened by polymerase chain reaction （PCR）-single strand conformation polymorphism （SSCP）/heteroduplex analysis （HA） and further confirmed by DNA sequencing. Results Among 28 families, three NF-1 patients （a father and 2 sons） came from one family with missense mutations for Leull41Arg of exon 20 T→G heterozygous mutation were found. At first, abnomal mobility shift was detected by SSCP/HA, when the same experiment was repeated, SSCP showed negative resuh and HA remained abnormal. In another NF-1 patient abnormal mobility shift was detected by SSCP/HA and heterozygous for G→T polymorphism located at-28 nueleotide of exon 28 was found in DNA sequencing. No mutation of exon 29, 39 was seen. Conclusion The combination of PCR-SSCP/HA can increase the sensitivity and positive detectable rate of NF1 gene mutation. Exon 20, 28, 29, 39 are not mutation hotspots or relative high mutation rate regions of NF1 gene in 56 Chinese patients from 28 examined families.