摘要
水茄果实中富含多糖、蛋白、色素和其他代谢物质,影响其总RNA提取的质量。本研究中比较Trizol法、改良Trizol法、改良CTAB法和两种试剂盒法5种方法提取的水茄果实总RNA,用琼脂糖凝胶电泳检测总RNA的完整性,Onedrop光谱仪检测总RNA的浓度和纯度。结果表明:改良Trizol法最为有效,该方法提取的总RNA在琼脂糖凝胶上得到清晰的28S和18S条带,OD260nm/OD280nm值在1.82.1之间、OD260nm/OD230nm的比值大于2.0;改良CTAB法获得的RNA质量较好,但步骤较为繁琐,耗时长;Trizol法和试剂盒法得到总RNA浓度较低。RT-PCR试验进一步表明,以改良Trizol法获得的总RNA可用于基因分离,所得条带清晰,与目的片段大小一致且比对正确。
Extracting total RNA from fruits of Solanum torvum Swartz. is difficult due to containing large amounts of polysaccharides, proteins, pigments and other metabolites, affecting the quality of total RNA. We compared the performances of five extraction methods, Trizol, modifiied Trizol, modifiied CTAB and two kinds of plant total RNA extraction kits in this research. The intrgrity of RNA were tested by agarose gel electrophoresis,the concentration and purity of RNA tested by One Drop spectrometer. The results showed that Modified Trizol method was the most efficient method, by which the total RNA isolated could be observed with two distinct electrophoresis bands of 28 S and 18 S r RNA with the OD_(260nm)/OD280 nm value ranging from 1.8 to 2.1, and the OD_(260nm)/OD_(230nm) value was more than 2.0. Total RNA was of good quality and successfully obtained by the method of Modified CTAB. However, the method was complicated and time exhausting. The total RNA obtained by Trizol and kit methods were of low concentration. RT-PCR analysis also showed that the extracted total RNA by modified Trizol method could be applicated on plant gene isolation, producing clear band, which was uniform with the size of target gene.
出处
《分子植物育种》
CAS
CSCD
北大核心
2016年第4期864-869,共6页
Molecular Plant Breeding
基金
江苏省自然科学青年基金项目(BK20140761)资助
