Recombinant strain of Rhodobacter sphaeroides as well as construction method and application of recombinant strain

Abstract

The invention relates to a recombinant strain of Rhodobacter sphaeroides as well as a construction method and an application of the recombinant strain. A recombinant strain with rshI gene knocked outis subjected to sdhB gene knockout, potassium tellurite and kanamycin are taken as single-crossover selection markers, compared with sdhB gene knockout of Rhodobacter sphaeroides with the original method, double-bacteria conjugative transfer efficiency is increased remarkably, conversion rate reaches 90% or higher and is increased 50-100 times as compared with a control group, and screening efficiency for markerless knockout of the Rhodobacter sphaeroides is increased remarkably; for the strain with the rshI gene knocked out, a selection marker is not needed to be inserted, the rshI gene is knocked out seamlessly at a fixed point, transcriptional level of upstream and downstream genes of the rshI gene is not affected, and normal growth of the Rhodobacter sphaeroides is guaranteed. Besides,an electron transport chain is modified in molecular biology means such as an efficient gene markerless knockout method of the Rhodobacter sphaeroides, and yield of coenzyme Q10 is increased by optimizing fermentation conditions.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to the rhodobacter sphaeroides recombinant bacteria and its construction method and application. Background technique [0002] Rhodobacter sphaeroides is a Gram-negative bacterium belonging to the α-proteobacteria group of purple non-comb bacteria. At present, the genetic modification of Rhodobacter sphaeroids mainly adopts conjugative transfer method. There are few reports that Rhodobacter sphaeroides cannot transfer plasmids by chemical transformation and electric shock transformation. At present, the efficiency of conjugative transfer of Rhodobacter sphaeroides to obtain positive transformants is generally low, so it is necessary to study the efficiency of conjugative transfer of Rhodobacter sphaeroides in order to improve the efficiency of genetic manipulation. It has been reported that the hsdR gene encoding the type I restriction endonuclease that degrades foreign DNA...

AI Technical Summary

Problems solved by technology

There are few reports that Rhodobacter sphaeroides cannot transfer plasmids by chemical transformation and electric shock transformation

At present, the efficiency of conjugative transfer of Rhodobacter sphaeroides to obtain positive transformants is generally low, so it is necessary to study the efficiency of conjugative transfer of Rhodobacter sphaeroides in order to improve the efficiency of genetic manipulation

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