摘要
目的研究HLA新的等位基因HLA A3308的分子机制。方法样本DNA抽提采用PEL FREEZ抽提试剂盒,应用PCR方法扩增先证者HLA A基因的第1～8外显子,PCR产物直接经TOPO转染克隆到质粒载体中获得等位基因的单链,对所得克隆进行第2、3、4外显子双向测序分析。结果先证者样本克隆测序得到两个等位基因,其中1个等位基因为A0201,另一个经BLAST验证其为新的等位基因,新的等位基因序列已递交GenBank(DQ089631,DQ089632,DQ089633)。与最接近的A3303等位基因序列相比,新的等位基因在第2外显子上有5个核苷酸不同,即第240位A→T,第256位C→G,第259位A→G,第261位C→G和第270位T→A;这导致3个氨基酸改变:第62位Arg→Gly、第63位Asn→Glu和第66位Asn→Lys。结论该等位基因为新的HLA A等位基因,被世界卫生组织HLA因子命名委员会正式命名为HLAA3308。
Objective To investigate the molecular genetics basis for HLA novel allele HLA-A ＊ 3308 in Chinese population, nethods DNA was extracted from whole blood by PEL-FREEZ DNA extraction kit. The amplification of HLA-A exons 1-8 of the proband was preformed by PCR and the amplified product was cloned with TOPO cloning sequencing kit to split the two alleles apart. Both strands of exons 2, 3 and 4 of chosen clones were sequenced. The PCRSSP was performed to confirm the mutations detected by sequencing. Results The sequencing results showed HLA-A alleles of the proband as A ＊ 0201 and the novel allele. The sequences of the novel allele have been submitted to GenBank （DQ089631, DQ089632, DQ089633）. BIAST analysis showed that the novel allele get the difference from A ＊ 3303 by five nucleotides at positions 240 A→T, 256C→G, 259A→G, 261C→G and 270T→A in exon 2. This resulted in three amino acids changes from Arg to Gly at cedon 62, Asn to Glu at cedon 63 and Asn to Lys at cedon 66. This allele is a novel allele and has been oflqcially named A ＊ 3308 by the WHO Nomenclature Committee.
出处
《中华医学遗传学杂志》
CAS
CSCD
北大核心
2006年第3期269-271,共3页
Chinese Journal of Medical Genetics
基金
浙江省医药卫生科学研究基金(2003Z003)~~
