Gene transient expression method with in-vivo jujube fruits as materials

Abstract

The invention relates to a gene transient expression method with in-vivo jujube fruits as materials. The method comprises the following steps of performing homologous cloning according to sequence information in an NCBI database to obtain a target gene, integrating the target gene into a plant overexpression plasmid vector through enzyme digestion-connection, and transferring a recombinant plant vector into agrobacterium GV3101; carrying out shake culture on the agrobacterium in an LB culture medium at 28 DEG C until OD600 reaches 1.0-1.2, centrifuging and collecting bacteria, pouring out theLB culture medium, and resuspending thalli by using a penetrating fluid containing MgCl2, MES and acetosyringone for later use; injecting the agrobacterium penetrating fluid into the in-vivo jujube fruits in the half-red period by using a medical injector; and verifying the function of the target gene by observing the phenotypic changes of the jujube fruits and detecting the expression quantity ofthe target gene and the content of related metabolites. The method avoids the problems of poor repeatability, low conversion rate, long time and the like of stable genetic transformation of ziziphusjujuba mill, can verify the gene function in a short time, is simple and quick, and saves the cost.

Description

technical field [0001] The invention belongs to the field of biotechnology engineering, and in particular relates to a gene transient expression method using in vivo jujube fruit as a material. Background technique [0002] jujube( Ziziphus jujuba Mill.) is Rhamnaceae (Rhamnaceae) jujube ( Ziziphus Mill.) plant, native to the middle and lower reaches of the Yellow River in my country, has been cultivated for more than 3,000 years. Since the reform and opening up, the jujube industry has developed rapidly and has become the largest dried fruit tree species in my country, accounting for 99% of the world's output. Jujube fruit is extremely nutritious. According to the measurement, the content of important therapeutic components such as cyclic adenosine monophosphate (cAMP), vitamin C, and carbohydrates in jujube fruit ranks among the top in all kinds of fruits. my country is in an international leading position in jujube research. In 2014, our research group took the lea...

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Problems solved by technology

[0004] The purpose of the present invention is to overcome the defect of prior art, provide a kind of gene transient expression method that takes body jujube fruit as material, utilize this method to directly carry out functional verification to numerous fruit development-related genes on jujube tree, avoid Many problems such as poor reproducibility, low conversion rate, and long time-consuming existing in the stable genetic transformation method of jujube were solved.

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