A New Pipeline Based on Target Capture Deep Sequencing Accurately Identifies Variants of α and β Globin Gene Clusters in Chinese Population
作者单位：Department of Medical GeneticsSchool of Basic Medical SciencesSouthern Medical University
会议名称：《第十四次全国医学遗传学学术会议》
会议日期：2015年
学科分类：0710[理学-生物学] 1002[医学-临床医学] 07[理学] 100201[医学-内科学(含：心血管病、血液病、呼吸系病、消化系病、内分泌与代谢病、肾病、风湿病、传染病)] 071007[理学-遗传学] 10[医学]
摘 要：Objective:Thalassemia is one of the most common monogenic disorders worldwide and it is most prevalent hemoglobinopathy in Southern *** detecting disease-causing variants is the basic step for further molecular and genetic *** we developed a new pipeline based on target capture deep sequencing that is able to identify all kinds of variants in hemoglobin gene ***:We selected559 thalassemia(including 22 carriers and 537 patients)samples and validated their genotypes of globin genes by laboratorial routine *** develop a reliable detection method based on Next-generation sequencing(NGS),we used customized NimbleGen array and lllumina HiSeq2500 2X100bp pair end indexing protocol to sequence the target *** quality control,mapping fastq reads with BWA and remove duplication with SAMTools,the pipeline is organized in three main steps:1)use properly mapped pair end reads to call SNPs and Indels with GATK,2)use unique mapped reads to discover CNVdepth and copy numbers with in-house python scrip and 3)Realign the unmapped reads against customized deletion reference database to identify the breakpoint-supporting split ***:A mean sequencing depth of 120-fold was achieved among the 559samples while these samples included 210 cases ofβ-thalassemia(14 heterozygotes,47 homozygotes and 196 compound heterozygotes),299 cases ofα-thalassemia(2non-deletional forms and 297 deletional forms which contains 288 HbH samples),and50 cases with disease-causing variants in bothαandβglobin *** accuracy rate is 100%in point mutations and Indels detection and 99.82%(558/559)in detecting large *** false negative result in that sample is probably due to the unstable capturing efficiency before sequencing ***:These data show that target capture deep sequencing is effective in exploring all the genetic variants especially for those rare mutations and novel ***,this method can also be applied to oth