cDNA分子克隆法鉴定KIR3DL3*064新等位基因  

Identification of a novel KIR3DL3*064 allele by cDNA cloning and sequencing

作　　者：聂冬梅[1] 蔡思齐 张国彬[1] 邓志辉[1] Nie Dongmei;Cai Siqi;Zhang Guobin;Deng Zhihui(Shenzhen Blood Center,Shenzhen,Guangdong 518035,China)

机构地区：[1]深圳市血液中心,518035

出　　处：《中华医学遗传学杂志》2020年第8期895-897,共3页Chinese Journal of Medical Genetics

基　　金：国家自然科学基金(81373158)。

摘　　要：目的:应用cDNA分子克隆测序方法,鉴定中国南方汉族1个KIR3DL3新等位基因。方法:采集1例KIR3DL3基因测序结果异常的无偿志愿捐血者的外周血样,提取mRNA并逆转录为cDNA。设计1对具有KIR3DL3基因特异性的PCR引物对cDNA进行扩增,扩增产物纯化后进行分子克隆测序。结果:经分子克隆测序,检出1个正常的KIR3DL3*00802等位基因和1个KIR3DL3*064新变异等位基因。KIR3DL3*064与KIR3DL3*00601等位基因的序列最为接近,仅有编码区存在nt1184 C>T错义变异,导致第9外显子中的第374密码子由ACT变为ATT,所编码的苏氨酸(Thr)变为异亮氨酸(Ile)。该等位基因被世界卫生组织HLA因子命名委员会KIR分委会正式命名为KIR3DL3*064。结论:cDNA分子克隆测序法能够鉴别常规测序分型中的异常结果,可用于鉴定KIR3DL3新等位基因。Objective To report on a novel KIR3DL3 allele identified in a southern Han Chinese individual.Methods Peripheral blood sample was collected from a voluntary blood donor with inconclusive result by KIR3DL3 sequence-based typing(SBT).Total mRNA was extracted and subjected to reverse transcription to obtain KIR3DL3 cDNA,which was then amplified by PCR with a pair of KIR3DL3-specific primers.The product was subjected to cDNA cloning and sequencing.Results cDNA cloning and sequencing have identified a wide-type KIR3DL3*00802 allele and a novel KIR3DL3*064 allele.The latter differed from KIR3DL3*00601 by a missense variant at codon 374[c.1184 C>T(p.Thr374Ile)]in exon 9.The novel KIR3DL3 allele has been officially assigned by the KIR subcommittee of World Health Organization Nomenclature Committee for factors of HLA system.Conclusion cDNA cloning and sequencing may be used to distinguish inconclusive results in KIR3DL3 SBT in order to identify novel KIR alleles.

关 键 词：KIR3DL3基因 CDNA 分子克隆测序 新等位基因 

分 类 号：R73[医药卫生—肿瘤]