海洋科学
海洋科學
해양과학
Marine Sciences
2015年
10期
8-14
,共7页
油九菊%范嗣刚%黄桂菊%郝博飞%陈飞飞%喻达辉
油九菊%範嗣剛%黃桂菊%郝博飛%陳飛飛%喻達輝
유구국%범사강%황계국%학박비%진비비%유체휘
合浦珠母贝(Pinctada fucata)%微卫星%磁珠富集
閤浦珠母貝(Pinctada fucata)%微衛星%磁珠富集
합포주모패(Pinctada fucata)%미위성%자주부집
Pinctada fucata%microsatellite%enrichment with magnetic beads
采用生物素标记的(AC)15探针和磁珠富集法构建合浦珠母贝(Pinctada fucata)基因组DNA微卫星富集文库。随机挑选2097个克隆进行筛选,得到483个候选克隆(23.03%),对其中135个阳性克隆测序分析发现122个克隆含有微卫星重复单元(90.37%)。进一步通过序列比对,最终得到65个具有特异微卫星序列的阳性克隆(53.28%),其中包含85个微卫星 DNA结构域,其中完美型(perfect)70个,占82.36%;非完美型(imperfect)7个,占8.24%;混合型(compound)8个,占9.41%,重复次数主要分布在5~20(95.74%),平均重复次数为7.83,(AC/GT)n重复所占比例最高(75.53%)。基于微卫星两端的侧翼序列,利用Primer Premier 5.0设计引物,获得11对具有多态性的微卫星引物。本研究为开展合浦珠母贝分子育种及资源评价分析提供了基础资料。
採用生物素標記的(AC)15探針和磁珠富集法構建閤浦珠母貝(Pinctada fucata)基因組DNA微衛星富集文庫。隨機挑選2097箇剋隆進行篩選,得到483箇候選剋隆(23.03%),對其中135箇暘性剋隆測序分析髮現122箇剋隆含有微衛星重複單元(90.37%)。進一步通過序列比對,最終得到65箇具有特異微衛星序列的暘性剋隆(53.28%),其中包含85箇微衛星 DNA結構域,其中完美型(perfect)70箇,佔82.36%;非完美型(imperfect)7箇,佔8.24%;混閤型(compound)8箇,佔9.41%,重複次數主要分佈在5~20(95.74%),平均重複次數為7.83,(AC/GT)n重複所佔比例最高(75.53%)。基于微衛星兩耑的側翼序列,利用Primer Premier 5.0設計引物,穫得11對具有多態性的微衛星引物。本研究為開展閤浦珠母貝分子育種及資源評價分析提供瞭基礎資料。
채용생물소표기적(AC)15탐침화자주부집법구건합포주모패(Pinctada fucata)기인조DNA미위성부집문고。수궤도선2097개극륭진행사선,득도483개후선극륭(23.03%),대기중135개양성극륭측서분석발현122개극륭함유미위성중복단원(90.37%)。진일보통과서렬비대,최종득도65개구유특이미위성서렬적양성극륭(53.28%),기중포함85개미위성 DNA결구역,기중완미형(perfect)70개,점82.36%;비완미형(imperfect)7개,점8.24%;혼합형(compound)8개,점9.41%,중복차수주요분포재5~20(95.74%),평균중복차수위7.83,(AC/GT)n중복소점비례최고(75.53%)。기우미위성량단적측익서렬,이용Primer Premier 5.0설계인물,획득11대구유다태성적미위성인물。본연구위개전합포주모패분자육충급자원평개분석제공료기출자료。
Microsatellite-enhanced genomic library of thePinctada fucata was constructed using a repeat-enrichment method with biotin-labeled oligos (AC)15 and streptavidin magnetic beads. From randomly selected 2097 clones, 483 clones (23.03%) containing microsatellite motifs were obtained by PCR screening. 135 positive clones were sequenced and 122 microsatellite loci (90.37%) were identified. By alignment, 65 microsatellite clones were unique. Sequence analyses of repeat motifs indicate that the microsatellites can be divided into three types: 70 perfect types (82.36%), 7 imperfect types (8.24%) and 8 compounds (9.41%). In addition, most microsatellite sequences con-tained 5-20 repeat units (95.74%) and the average was 7.83. The proportion of (AC/GT)n repeat was the highest (75.53%). Based on the flanking sequences at both ends of the microsatellite, 11 pairs of microsatellite polymorphic primers were obtained at last, which were effective for PCR amplification inP. fucata genome. This study provides a base for molecular assistant selective breeding and assessment of germplasm resources ofP. fucata.