HLA新等位基因A*9217家系调查分析

Investigation for the heritage of a new HLA allele A*9217

背景:目前所发现的HLA等位基因是在人类进化过程中,为了适应某种外界环境过程中逐渐产生的,新近产生而不曾被人们发现的为新等位基因,这些新的等位基因产生诱因千变万化,在诱发因素消除之后这些突变是否持续存在,尚需进一步观察。目的:采用DNA测序技术确认聚合酶链反应-序列特异性寡核苷酸分型检测中的可疑结果,确认新的HLA等位基因,分析新等位基因HLA-A*9217(WHO注册号:WHS10004629)的遗传方式。设计、时间及地点:以DNA为观察对象的开放性试验。其中常规初检聚合酶链反应-寡核苷酸探针分型反向杂交荧光微珠法于2007-11在河南省红十字血液中心中华骨髓库河南分库人类白细胞抗原组织配型实验室完成,测序实验于2008-02在戴诺生物技术(北京)有限公司人类白细胞抗原实验室完成。材料:中华骨髓库志愿捐献者(编号:371xxxxx)及其直系3代亲属血样,受试者共9人在实验室逐个登记,采集静脉血5mL,EDTA抗凝。方法:用聚合酶链反应-序列特异性寡核苷酸流式荧光微珠HLA分型方法、SBT测序分型方法,对A*9217携带者家庭成员进行中低分辨,DNA测序高分辨检测分析,血清学方法检测ABO,MN,Rh血型系统,分析新基因的发现及遗传方式。主要观察指标:HLA新等位基因A*9217携带者A位点外显子3序列对比。结果:通过受检者HLA分型结果和红细胞系统ABO,MN,Rh血型分析,先证者A*9217应来自父方,但父方并没有检出A*9217,而是检出了与之序列相近的A*020301,与A*9217在处显子3有3处碱基改变391T>G,414C>G,418T>G,A*9217携带者的2个孩子均检出A*9217。结论:新基因A*9217先证者父方产生突变所致,该突变能正常遗传给后代,该基因是否能稳定遗传下去有待进一步观察研究。

BACKGROUND：The human leucocyte antigen （HLA） allele are now understood to be alternative DNA sequences at the same physical gene locus,which may or may not result in different phenotypic traits.The generation of a new allele is induced by various factors,but,whether the mutation would be exist after removing the causative factors need further investigation.OBJECTIVE：To confirm the new allele by DNA sequencing technique,in addition,to analyze the heritage of a HLA allele A＊9217 （No.of WHO registration：WHS10004629） DESIGN,TIME AND SETTING：The open experiment with DNA as observation object.The initial detection with polymerase chain reaction sequence-specific oligonucleotide （PCR-SSO） was performed at HLA Laboratory of Henan Provincial Red Cross Blood Center in November 2007,and the sequencing was performed at HLA Laboratory of DYNAL Biological Technology （Beijing） Co.,Ltd.in February,2008.MATERIALS：The proband （sample ID：371xxxxx） and other 8 family member was investigated,blood sample was collected at the HLA Laboratory.METHODS：Nine family members of A＊9217 carrier were typing for HLA-A,B,DRB1 using PCR-SSO and SBT methods for low-media and high resolution,and 3 red blood cell blood groups systems：ABO,MN and Rh was tested to assist analysis.MAIN OUTCOME MEASURES：Sequence alignment of HLA-A exon 3 in A＊9217 carriers.RESULTS：According to the results of the blood groups phonotype of ABO,MN,Rh systems and HLA,the new allele A＊9217 of the proband was paternal origin.However,the sequence of this allele is A＊020301,which had 3 base difference in exon 3,the new allele appears 3 base change at 391 TG,414 CG,418 TG,and this new sequence was found in proband＇s two children.CONCLUSION：The new allele A＊9217 is generated from proband＇s father mutation,which can pass it to his children.However,the further heritage of A＊9217 allele need exploring.