| Paenibacillus polymyxa SC2 is a plant rhizosphere-promoting bacteria selected from pepper rhizosphere soil.It has a broad antibacterial spectrum and has obvious growth-promoting effects on various crops.Showed an unstable state,and two spontaneous mutant strains SC2-M1 and SC2-M2 were produced.The colony of strain SC2-M1 is irregular in shape,transparent in color,low in extracellular polysaccharides,does not produce spores,and is easy to be electrotransformed.In this experiment,the mutant strain SC2-M1 was used as the background strain,and the functions of the genes lacF3 and yvqE1 were analyzed.From previous genome-wide sequencing,we found that the mutant strain SC2-M1 had multiple single nucleotide mutations compared to the wild-type strain SC2,among which lacF3 and yvqE1 are two important mutant genes.In this experiment,the function of genes was mainly analyzed by achieving overexpression of lacF3 and yvqE1 in mutant strain SC2-M1.Using the pHY300 PLK plasmid and tetracycline as resistance screening markers,lacF3 and yvqE1 overexpression vectors were constructed,and the constructed overexpression vectors were transformed into SC2-M1 by electric transformation to obtain the lacF3 gene overexpression strains LM1 and yvqE1 gene overexpression strains yvM1.At the same time,the empty plasmid pHY300 PLK was electrotransformed into SC2-M1 in the same way to obtain the control strain p M1.The over-expressing strain yvM1 was mainly studied for tolerance to abiotic stress,including the determination of heat stress,osmotic stress,hydrochloric acid stress,and the determination of acid production capacity.The results showed that the growth of the over-expressing strain yvM1 was higher than that of the control strain p M1 under several different stress environments,and the acid-producing ability of the over-expressing strain was stronger than that of the control strain in different carbon source fermentation media.It is speculated that the yvqE1 gene regulates the acid produced by sugar fermentation and plays an important role in strains responding to various environmental stresses.The carbon source utilization ability of the over-expressing strain LM1 was determined,that is,eight sugars were selected as the sole carbon source in the fermentation medium to determine the growth ability.The results showed that the growth of the over-expressing strain LM1 in different carbon sources was higher than that of the control p M1,demonstrating that the lacF3 gene had different effects on the growth of the strain.In order to further study the effect of lacF3 gene on the metabolism of the bacteria,we used transcriptome sequencing and metabonomics analysis to study the gene expression difference between LM1 and PM1 under the same condition,so as to obtain the relevant information.The results of transcriptome sequencing showed that there were 3625 differential genes in LM1 and p M1,of which 2023 were up-regulated and 1602 were down regulated.Among them,there were more differential metabolism genes related to carbohydrate metabolism,and some of them were up-regulated in PTS and ABC.A metabolome analysis revealed a total of 169 differential metabolites,of which 83 were up-regulated in the positive ion mode,38 were down-regulated,and 31 were up-regulated and17 down-regulated in the negative ion mode.By analyzing the metabolic pathway information of differential metabolites,it was found that the detected metabolites participate in multiple metabolic pathways,including the tricarboxylic acid cycle(TCA)cycle,starch and sucrose metabolism pathways,galactose metabolism pathways,and phosphotransferase system(PTS),amino acid metabolism pathway,purine metabolism pathway,which are consistent with transcriptome sequencing information.The results of this research have initially demonstrated the related functions of the lacF3 and yvqE1 genes,laying a foundation for further elucidating their related molecular mechanisms. |
PDF Full Text Request