摘要
柴胡是我国传统中药材,主要用于治疗感冒及保肝护肝等。三岛柴胡是日本栽培品种,在我国和韩国等国家地区亦有种植。在大规模全基因组深度测序之前,需要进行低覆盖度的基因组Survey测序,以评价基因组的大小及复杂程度,确定适合该植物全基因组测序研究策略。该研究采用二代高通量测序技术(Illumina Hiseq 2000)进行了三岛柴胡的Survey测序分析,并对所获得基因组序列进行了SSR分析,利用Primer 3设计SSR特异引物并随机选择33对引物以三岛柴胡DNA为模板,进行PCR扩增,对PCR体系以及最佳退火温度进行筛选。共获得了288.64 G基因组数据,估计基因组大小为2119.58Mb,测得基因组数据深度为138×,杂合率为1.84%,重复序列比例为83.89%,推测三岛柴胡基因组属于复杂基因组。采用K-mer=41初步组装,得到contig N50224 bp,总长896.97 Mb,scaffold N50313 bp,总长922.67 Mb。三岛柴胡基因组序列数据中,共检测到91377条SSR序列,分布于70809条独立基因。主要类型为二核苷酸重复,存在49680条,占比70.16%。在合成的33对引物中,有21对引物成功扩增出目标条带。研究结果为后续大规模基因组测序、开发鉴定柴胡种质和性状定位的SSR分子标记奠定了基础。
Buplewrum falcatum is a traditional Chinese medicine,which is mainly used for the treatment of cold and liver protection.B.falcatum is dominantly cultivated in Japan as well as planted in China,Korea and other countries and regions.In order to determine the appropriate sequencing strategy,the genome survey before large-scale genome sequencing is needed.This survey can provide information about the size and complexity of the whole genome of the target species.In the present study,the next generation sequencing technology(Illumina Hiseq 2000)was used to analyze the genome size and complexity of B.falcatum.In addition,SSR loci were analyzed from the sequenced data.Primer 3 was used to design specific primers and 33 pairs of primers were randomly selected for PCR with template DNA of B.falcatum,and the PCR system and optimal annealing temperature were screened.A total of 288.64 G genome sequence data was obtained,and the estimated genome size of B.falcatum was 2119.58 Mb.The measured genome data depth was138×;the rate of heterozygosity was 1.84%;and the ratio of repeat sequence was 83.89%.It is speculated that the genome of B.falcatum is complex.The preliminary assembly was performed with K-mer=41,and the contig N50 was 224 bp,the total length 896.97 Mb,the scaffold N50313 bp,and the total length was 922.67 Mb.A total of 91377 SSR sequences were detected in the sequenced genome data which were distributed in 70809 unigenes.The main type is dinucleotide repeats,with 49680 sequences,accounting for70.16%.Among the 33 pairs of primers randomly synthesized according to the obtained SSR sequences,21 pairs were successfully amplifying the target sequences.The results will be helpful for later large scale genome sequencing and SSR molecular markers development for germplasm identification and trait mapping.
作者
朱楚然
徐娇
都明理
王丽红
隋春
魏建和
ZHU Chu-ran;XU Jiao;DU Ming-li;WANG Li-hong;SUI Chun;WEI Jian-he(Institute of Medicinal Plant Development(IMPLAD),Chinese Academy of Medical Sciences&Peking Union Medical College(Key Laboratory of Bioactive Substances and Resources Utilization of Chinese Herbal Medicine,Ministry of Education&National Engineering Laboratory for Breeding of Endangered Medicinal Materials),Beijing 100193,China;Jiamusi University,Jiamusi 154007,China)
出处
《中国中药杂志》
CAS
CSCD
北大核心
2019年第18期3960-3966,共7页
China Journal of Chinese Materia Medica
基金
四川省科技支撑计划项目(2016NYZ0020)
中国医学科学院医学与健康科技创新工程(重大协同创新项目)(2016-I2M-2-003)
作者简介
通信作者:隋春,E-mail:csui@implad.ac.cn;通信作者:魏建和,E-mail:wjianh@263.net;朱楚然,E-mail:840637561@qq.com。
