Method for improving instantaneous expression rate of agrobacterium tumefaciens-mediated sugarcane glucuronidase (GUS)

Abstract

The invention relates to a method for improving the instantaneous expression rate of an agrobacterium tumefaciens-mediated sugarcane glucuronidase (GUS). The method comprises the following steps of: selecting materials, preparing conversion bacteria liquid, performing an infection method, performing cocultivation and selective cultivation, and chemically dyeing GUS active tissues. The method that the advantages and disadvantages of a transgenic technology method is measured by taking the degree of the GUS instantaneous expression rate as an index is nationally acknowledged. Therefore, the index which is nationally acknowledged is adopted to evaluate the technology method. By the method, the GUS instantaneous expression rate can be increased by 335.8 to 1,859.3 percent, the bottle neck problem about efficiently obtaining a transgenic sugarcane plant by using a agrobacterium tumefaciens-mediated way can be effectively solved, a key technology method for introducing an exogenous gene into a sugarcane is provided for improving the variety of the sugarcane by using transgenic technology. Meanwhile, the method is also applied to the genetic transformation for other plants based on an agrobacterium tumefaciens-mediated way.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to a method for improving the transient expression rate of GUS in sugarcane mediated by Agrobacterium. Background technique [0002] sugar cane( Saccharum Complex) is the most important sugar crop. Sucrose accounts for 76% of the world's total sugar production and 92% of my country's total sugar production. Sugarcane is also one of the most important biomass energy crops. C4 plants with the largest amount and fuel ethanol production. The contribution rate of sugarcane variety improvement to industrial scientific and technological progress is as high as 60%, but due to the large genome of sugarcane, the genetic background is very complex, the number of chromosomes is more than 120, and the modern sugarcane cultivated species is the interspecific hybridization of 3 or 4 species of Succulus The complex belongs to allopolyploid plants. At the same time, the target object of sug...

AI Technical Summary

Problems solved by technology

There are mainly two approaches to transgenic technology in a narrow sense, one is the gene gun bombardment method, and the other is the Agrobacterium-mediated method. The former has many application examples in sugarcane, but this technology has its inherent shortcomings. In addition to expensive equipment, high operating costs, and uncertain target gene source insertion sites, almost all transgenic lines obtained by this technology have multiple copies, which is not conducive to the stable inheritance of traits and transgenic safety, and is prone to gene silencing. The imported exogenous gene is easy to lose and other problems; the insertion site of the latter is clear, the frequency of obtaining single-copy transgenic offspring is high, and the genetic stability of traits is high. relatively high

Although the Agrobacterium-mediated method for several monocotyledonous plants such as rice has been relatively mature, compared with dicotyledonous plants, the Agrobacterium-mediated transformation rate of monocotyledonous plants is generally relatively low, and Agrobacterium-mediated transformation of sugarcane The efficiency is still relatively low, and the improvement of transformation efficiency is the "bottleneck" problem of sugarcane transgenic breeding, because only a certain number of transgenic sugarcane populations can be screened for excellent single plants

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