RNAi (RNA interference)-based transgenic novel watermelon material construction method

Abstract

The invention relates to an RNAi (RNA interference) vector construction method and an application thereof in watermelon genetic transformation, relates to the technical field of plant virus control and belongs to the application technology in biological and modern agriculture technique fields. The mediated virus infection function is destroyed by acting on host factors and thus watermelons obtain antiviral activity, which has not been reported yet at present. An RNAi vector is constructed and genetically transformed to watermelons, the constructionmethod is characterized in that watermelon eIF4E and a homological isomer eIF(iso)4E gene thereof are cloned and connected, 4E-iso sense and antisense fragments are obtained and connected to two sides of an intron sequence, and a 4E-iso hairpin structure fragment is obtained; the 4E-iso sense fragment, the 4E-iso antisense fragment and the hairpin structure fragment are inserted into a pCAMBIA1301 vector through T4 connection and homologous recombination, and the RNAi vector is obtained; target gene fragments are transformed into watermelon cotyledon explants through agrobacterium tumefaciens-medicated transformation, and transgenic watermelons with the target gene fragments are obtained through screening cultivation. The method is mainly used for providing a novel material and a novel technology for watermelon virus and disease control.

Description

technical field

[0001] The invention belongs to the technical field of molecular biology, and in particular relates to the construction of an RNA interference vector suitable for genetic transformation of dicotyledons and highly resistant to watermelon mosaic virus disease and its application in genetic transformation of watermelon. It further relates to the expression vector of the watermelon eIF4E gene and its homologous isomer eIF(iso)4E gene and the genetic transformation of the watermelon to obtain transgenic plants. technical background

[0002] The cultivation area of ​​watermelon (Citrullus lanatus) in my country ranks first in the world, with an average annual cultivation area of ​​1.2 million ha 2 . Watermelon virus diseases represented by watermelon mosaic virus (WMV) are the main limiting factors for the production of cucurbit crops such as watermelon, and the annual incidence of watermelon virus diseases reaches 100 thousand hm 2 , because of its serious harm,...

Images