摘要
不同PCR引物对有孔虫DNA扩增效能存在显著差异,可能导致PCR扩增的效率降低或失败。但目前为止,尚无对于不同海域生境沉积物类型(例如潮间带、近岸浅水或深海沉积物)的有孔虫引物进行有效DNA扩增方法学的比较研究。本实验有孔虫样品采自青岛湾、胶州湾、黄海、西北太平洋以及东太平洋海域表层沉积物,通过使用不同的PCR特异性引物对有孔虫SSU rDNA基因进行PCR扩增,比较不同引物获取的PCR产物的凝胶电泳结果,利用Image J软件对电泳图中目的条带的亮度进行定量分析,研究了不同扩增引物对多种沉积物类型中有孔虫SSU rDNA分子目的片段的扩增效能差异。结果显示,对于潮间带沉积物类型,s14F3和s17是扩增底栖有孔虫的有效组合;对于近岸浅海沉积物,s14F1和s17是有效组合;对于离岸陆架沉积物,使用s14F3和s15ROTEX有效;对于深海泥质沉积物,使用s14F3和s17较为有效;对于金属结核区沉积类型沉积物,使用s14F1和s15ROTEX更为有效。研究表明,由于不同水深沉积物、沉积物自身特性以及有孔虫种类组成存在差异,扩增有孔虫DNA应该根据不同的沉积物类型选取合适的PCR引物,以提高有孔虫遗传物质的扩增效能。本研究结果可为涉及不同海域沉积物类型的底栖有孔虫分子多样性等研究提供有效的技术方法参考。
The amplification efficiency of foraminifera DNA differs based on different PCR primers.The improper primers can even cause the failure of PCR amplification.However,it has no idea that which foraminiferal primer is effective on DNA amplification in different sediment types of diverse marine habitats(intertidal zone,coastal area or deep sea).Here,a part of the foraminiferal 18 S SSU rDNA gene of environmental genome(eDNA)collected from the surface sediments of Qingdao Bay,Jiaozhou Bay,Yellow Sea,Northwest Pacific and East Pacific Ocean was amplified used different foraminifera-specific primers.We used Image J software to quantize the gel electrophoresis results of PCR target bands obtained from different primers and compared the efficiency of different primers to the foraminiferal DNA from different sediments.The results showed that s14 F3(forward primer)and s17(reverse primer)were effective for intertidal sediments;and s14 F1(forward primer)and s17 were more effective for shallow coastal sediments;s14 F3 and s15 ROTEX(reverse primer)had a better amplification effect on the shelf sediments;and s14 F3 and s17 exhibited better effects for multi-deep-sea muddy sediments;s14 F1 and s15 ROTEX were effective for multi-metal nodule deposits.In addition,it indicated that the water depth of sediment and the sediment itself(biological and non-biological)as well as the composition of foraminifera in the sediments had an important effect on the amplification efficiency of different primers.Thus,the appropriate primers would improve the amplification efficiency of PCR for foraminifer from different sediments.Our study will provide an effective technical method and reference basis for studies involving the molecular diversity of benthic foraminifera in different marine sediment types.
作者
沈阳阳
类彦立
李浩天
石峻峰
李青霞
曹一飞
SHEN Yangyang;LEI Yanli;LI Haotian;SHI Junfeng;LI Qingxia;CAO Yifei(Laboratory of Marine Organism Taxonomy and Phylogeny,Institute of Oceanology,Chinese Academy of Sciences,Qingdao26607;University of Chinese Academy of Sciences,Beijing100049;Center for Ocean Mega-Science,Chinese Academy of Sciences,Qingdao266071)
出处
《微体古生物学报》
CSCD
北大核心
2020年第4期368-380,共13页
Acta Micropalaeontologica Sinica
基金
国家自然科学基金项目(No.41976058
U1906211)
东太平洋生态环境监测保护(DY135-E2-5)
“全球变化与海气相互作用”专项“亚洲大陆边缘的古海洋与古地理演化”任务(GASI-GEOGE-04)
泰山学者工程专项(批准号:TS20190963)的支持。
关键词
底栖有孔虫
潮间带
近岸浅水
离岸陆架
深远海
分子生物学
方法学
benthic foraminifera
intertidal zone
shallow nearshore water
shelf
deep sea
molecular biology
methodology
