苹果MicroRNA的茎环RT-PCR检测及离体试管苗与田间苗表达差异

Detection of MicroRNA in Apple by Stem-loop RT-PCR and Different Expression of Them between in Vitro Plants and in Vivo Plants

MicroRNA(miRNA)在植物的生长发育过程中起着重要作用。以改进CTAB法提取的总RNA为模板,通过设计茎环反转录引物,利用定性RT-PCR技术从寒富苹果幼嫩叶片、成熟叶片、嫩茎、幼果和根等不同器官中均检测到miRNA,并从寒富苹果叶片中克隆分离出14种miRNA。在此基础上利用半定量RT-PCR技术对寒富苹果离体试管苗与田间苗中10种miRNA的表达差异进行分析,结果发现,miR156和miR157在离体试管苗叶片中表达量明显高于田间苗幼叶。本研究建立了苹果miRNA的茎环RT-PCR检测体系,为研究miRNA在苹果植株生长发育过程中的作用奠定了基础。

MicroRNA（miRNA） play an important role in plant growth and development.Using total RNA extracted by modified CTAB method as template,miRNA were detected by qualitative RT-PCR with stem-loop RT primers in young leave,mature leave,tender stem,young fruit and root of Hanfu apple.Fourteen kinds of miRNA sequences were identified in leaves of Hanfu apple.Semi-quantitative RT-PCR was used to investigate the expression difference of ten miRNA between in vitro plants and in vivo plants of Hanfu apple.It was found that the expressions of miR156 and miR157 were obviously upregulated in the young leaves of in vitro plants.The system of analyzing expression of miRNA by stem-loop RT-PCR in apple plants laid a sound foundation for further studying the effect of miRNA in apple growth and development.