Method for rapidly and efficiently screening rhamnolipid producing bacteria nutrition system

Abstract

The invention relates to a method for rapidly and efficiently screening a rhamnolipid producing bacteria nutrition system. The method comprises the following steps: synthesizing a primer of rhamnolipid producing gene rhlAB, performing gene amplification by using PCR, synthesizing rhlAB gene fragments, connecting the rhlAB gene fragment to the position in front of lux luminescence gene on pMS402 plasmid by utilizing gene engineering shearing and splicing technology, then transferring the recombinant plasmid into pseudomonas aeruginosa DN1, coating the recombinant strain onto an LB solid plate containing 200mg/L-300mg/LTmp, screening out positive clone, naming the strain as rhlAB-lux strain, picking out positive monoclonal colonies to be enriched on the solid LB culture medium, refrigeratingat at 4DEG C for later use; taking out the refrigerated rhlAB-lux strain, and enriching the thallus overnight; and screening a nutrient optimized system. According to the method, the luminescence value can be determined by a multifunctional microplate reader, and the method has the characteristics that the flux is high, and the screening process is simplified and rapid.

Description

technical field [0001] The invention relates to a method for rapidly and efficiently screening the nutritional system of rhamnolipid-producing bacteria. Background technique [0002] In recent years, marine oil spills have emerged one after another, which has brought serious pollution problems to the marine and coastal environments. Oil spill pollution has caused a large number of biological deaths. Therefore, it is very important to understand how to use microorganisms to degrade hydrocarbons. Although hydrocarbons are relatively stable molecules, over millions of years, with the evolution of organisms, hydrocarbons have been degraded and utilized by microorganisms as the only carbon source and energy source. According to literature reports, at least 175 species of Microorganisms can degrade and utilize hydrocarbons. The main pollutants in hydrocarbons are aromatic hydrocarbons. Low-molecular-weight PAHs are easily degraded and utilized by various microorganisms, and high-...

AI Technical Summary

Problems solved by technology

The preparation of rhamnolipids mainly adopts microbial fermentation method. The fermentation raw materials of this process are widely and cheap, safe and economical, the process is simple, the product can be completely biodegraded, and it is not harmful to organisms and the environment. There are still some problems and defects in the production and development of lipids

From the perspective of the fermentation process itself, the production has relatively high production costs and low yields, and microbial strains are usually pathogenic or difficult to handle on a large scale, and the products produced are all mixtures, which cannot be achieved as a single product. Therefore, the downstream processing cost is increased. As far as the fermentation strain itself is concerned, the product yield is low, and the requirements for the medium and adaptability are weak. According to literature reports, if certain fermentation conditions are limited, the production of rhamnolipids can be increased. Yield, therefore, if a kind of nutrient optimization system for rapidly determining high-yielding rhamnolipids of Pseudomonas aeruginosa can be developed based on this principle, a lot of work will be saved

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