关键词: MicroRNAs, RT-PCR, 草莓, 鉴定

Abstract:

【Objective】 microRNAs (miRNAs) play an important role in plant growth and development. The aim of this paper is to develop a quick method for exact identification of the conserved miRNAs from plants. 【Method】 High quality small RNAs (less than 150 bases) were isolated from strawberry leaves with the modified CTAB method. Three kinds of RT-PCR methods, using 3′ adaptor, using 5′ and 3′ adaptors and stem-loop primers, were compared. Three kinds of templates, total nucleic acid, total RNA and small RNAs were compared in identification of miRNAs with stem-loop primers. 【Result】 For twenty miRNAs of plants, seven miRNAs were identified from strawberry plants by using 3′ adaptor, only one miRNA was successfully identified with the method of 5′ adaptor and 3′ adaptor, while fifteen miRNAs were identified by using stem-loop primers. No difference was found in PCR fragments by using three kinds of nucleic acids as templates. 【Conclusion】 Because the extraction of total nucleic acid was very simple and rapid, the suitable method for identification of the conserved miRNAs from plants with RT-PCR was to use stem-loop primers and total nucleic acid as template. This method had been approved on other plants, such as apple and grape.

Key words: microRNAs, RT-PCR, strawberry, identification