Method for constructing fluorescence labeled rhizobia

Abstract

The invention discloses a method for improving conjugational transfer efficiency of fluorescent plasmid transfer of fluorescence labeling strains of rhizobia by taking sterilizing alfalfa leaves as triparental mating carriers instead of sterile filter membranes. The method can be used for efficiently constructing CFP (cyan fluorescent protein) gene labeled strains of rhizobia. The method specifically comprises the following steps of taking small and tender leaves of alfalfa, uniformly putting the leaves in a single layer in a culture dish after washing the leaves with distilled water, and sterilizing the leaves; taking out the leaves with a pair of sterile tweezers, sticking the leaves to the surface of TY culture media, and covering the culture dish; and using the leaves to replace microporous filter membranes in the triparental mating process to carry out mixed growing culture on donors, assistant bacteria and recipient bacteria. Compared with the microporous filter membranes as the mating carriers, the alfalfa leaves, as the mating carriers, can improve the conjugational transfer efficiency of the fluorescent plasmids by 8.4-11 times, and the difference is obvious. As the used raw materials are alfalfa plant leaves which are renewable and accessible, the method has the advantage that the raw materials are cheap and accessible, is simple and convenient to operate and is suitable for constructing the fluorescence labeling strains of rhizobia.

Description

technical field [0001] The invention relates to the field of crop inoculants, in particular to a method for constructing fluorescently labeled rhizobia. Background technique [0002] Rhizobia inoculants are commonly used in agricultural production, and the yield of leguminous crops has also been significantly increased due to the application of rhizobia inoculants. also needs to be resolved. The development of rhizobia labeling technology and its application in scientific research and practical production have gradually clarified the nitrogen fixation mechanism of rhizobia and the process of infection and colonization in the roots of leguminous plants. One of the effective methods of competition and interaction with other microorganisms in the body. At present, the labeling effect is the best, and the easiest and quickest method for determination is fluorescent protein labeling of rhizobia. The CFP (Cyan Fluorescent Protein) gene is a mutant extracted from the jellyfish g...

AI Technical Summary

Problems solved by technology

However, due to the artificially prepared single-layer filter membrane structure, the nutrients in the medium can only be absorbed by the cells through the micropores, and the proliferation rate and conjugation transfer rate of the bacteria are limited.