Objective To explore the accuracy and the feasibility of this method in clinical application,single molecule real-time(SMRT)sequencing technology was employed to determine HLA genotyping,haploid type.Methods Blood samples were collected from 19 patients who needed kidney transplantation,genomic DNA was extracted respectively,and the sequences of all samples were obtained by SMRT sequencing technology,and then detected by PCR-SBT to verify the accuracy of the result.Results The genotypes and haplotypes of 19 samples could be assigned by SMRT technique,which were consistent with those of the PCR-SBT method.The results of HLA-C locus showed ambiguous combination on samples 1 and 14 using SBT method,with HLA-C*03:02,03:03 and-C*03:04,03:132;while the unique result of them is HLA-C*03:02:02,03:03:01 using SMRT technology.In addition,a new HLA-B allele of sample 1 was identified using SMRT method.Conclusion SMRT sequencing technology has high accuracy when applied to HLA high-resolution typing,and has unique advantages of accurately locating multiple long-distance mutation in HLA haploid typing.
关键词
单分子实时测序/长读长测序/HLA分型/单体型分析
Key words
Single molecule real-time sequencing/Long read sequencing/HLA typing/Haploid typing
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