Whole Genome Analysis of Bradyrhizobium sp. Bd1 and the Negative Regulating Function of TetR3 during Cell Growth and Nodulation

doi:10.13560/j.cnki.biotech.bull.1985.2025-0215

Abstract

Abstract:

Objective This research is aimed to understand the genetic background of Bradyrhizobium sp. Bd1 at the genome level, and explore effective ways to promote the growth and nodules of Bd1. Method Illumina + PacBio third-generation sequencing platform was used to sequence the whole genome of Bd1. Bioinformatics method was employed to analyze the species identification, functional gene annotation, nitrogen fixation-related genes and TetR family transcription factors. TetR3 gene knockout mutant was constructed by homologous recombination double-exchange method to investigate the function of TetR3 in the growth and nodulation of Bd1. Result The genomic average nucleotide consistency (ANI) and digital DNA-DNA hybridization values between Bd1 and Bradyrhizobium diazoefficiens USDA110 were 99.98% and 98.7%, respectively. Bd1 genome size was 9 009 469 bp with 64.1% guanine-cytosine content, 8 403 coding sequences, within them 51 tRNA coding genes and 3 sets of ribosomal RNA genes. The number of nod, nif and fix genes were 9, 12 and 10, respectively. There were 58 genes encoding TetR family transcription factors with diverse structure. Compared with the wild Bd1, the growth and nodulation capacity were significantly enhanced, and glutathione S-transferase activity increased by 41.0% in TetR3-inactive mutant (Bd1ΔTetR3). Conclusion Bd1 is a novel strain belonging to B. diazoefficiens, which has the ability of nitrogen fixation in symbiotic nodulation with soybean. TetR3 negatively regulates the growth and nodulation in Bd1 by balancing cell REDOX capacity.

Key words: Bradyrhizobium, whole genome sequencing, TetR, soybeans, glutathione S-transferase

LI Ya-tao, ZHANG Zhi-peng, ZHAO Meng-yao, LYU Zhen, GAN Tian, WEI Hao, WU Shu-feng, MA Yu-chao. Whole Genome Analysis of Bradyrhizobium sp. Bd1 and the Negative Regulating Function of TetR3 during Cell Growth and Nodulation[J]. Biotechnology Bulletin, 2025, 41(9): 289-301.

Figures/Tables 8

References 32

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Family	Gene ID	Name	KEGG gene ID	KO description	Location
fix	gene5832	fixK	bja：bll3466	CRP/FNR family transcriptional regulator	6070103-6070786
	gene6542	fixK	bja：bll2757	CRP/FNR family transcriptional regulator	6868843-6869541
	gene6539	fixL	bja：bll2760	LuxR family， sensor kinase FixL	6866141-6867658
	gene6540	fixJ	bja：bll2759	LuxR family， response regulator FixJ	6867662-6868279
	gene7359	fixA	bja：blr2038	Flavoprotein beta subunit	7803669-7804535
	gene7895	fixA	bja：blr1377	Flavoprotein beta subunit	8425108-8425857^C
	gene7566	fixX	bja：bsr1775	Ferredoxin like protein	8067228-8067524^C
	gene7567	fixC	bja：blr1774	Flavoprotein-quinone oxidoreductase	8067563-8068870^C
	gene7568	fixB	bja：blr1773	Flavoprotein alpha subunit	8068882-8069991^C
	gene7894	fixB	bja：blr1378	Flavoprotein alpha subunit	8424164-8425108^C
nod	gene5597	nodT	bsym：CIT39_22430	Multidrug efflux system	5805752-5807308
	gene7363	nodU	bja：blr2034	Carbamoyltransferase	7809897-7811504^C
	gene7367	nodU	bjp：RN69_37980	Carbamoyltransferase	7813893-7815602^C
	gene7365	nodJ	bja：blr2031	Lipooligosaccharide transport system permease	7812179-7812967^C
	gene7366	nodI	bja：blr2030	Lipooligosaccharide transport system ATP-binding protein	7812971-7813891^C
	gene7368	nodC	bja：blr2027	N-acetylglucosaminyltransferase	7816172-7817629^C
	gene7369	nodB	bja：blr2026	Chitooligosaccharide deacetylase	7817644-7818303^C
	gene7370	nodA	bja：blr2025	Nodulation protein A	7818300-7818932^C
	gene7372	nodD	bja：bll2023	LysR family transcriptional regulator	7819726-7820670
nif	gene7360	nifA	bja：blr2037	Nif-specific regulatory protein	7805053-7806801^C
	gene7569	nifW	bja：blr1771	Nitrogenase-stabilizing/protective protein	8070942-8071283^C
	gene7571	nifQ	bju：BJ6T_80490	Nitrogen fixation protein NifQ	8071938-8072714^C
	gene7572	nifH	bja：blr1769	Nitrogenase iron protein NifH	8072861-8073745
	gene7580	nifZ	bjp：RN69_39140	Nitrogen fixation protein NifZ	8078154-8078468^C
	gene7582	nifB	bju：BJ6T_80610	Nitrogen fixation protein NifB	8078954-8080537^C
	gene7583	nifT	bja：bsr1757	Nitrogen fixation protein NifT	8081201-8081425^C
	gene7594	nifX	bja：blr1747	Nitrogen fixation protein NifX	8090471-8090866^C
	gene7595	nifN	bja：blr1746	Nitrogenase molybdenum-iron protein NifN	8090863-8092272^C
	gene7596	nifE	bja：blr1745	Nitrogenase molybdenum-cofactor synthesis protein NifE	8092282-8093925^C
	gene7597	nifK	bja：blr1744	Molybdenum-iron protein beta chain	8094018-8095574^C
	gene7598	nifD	bja：blr1743	Molybdenum-iron protein alpha chain	8095640-8097142^C

Family	Gene ID	Name	KEGG gene ID	KO description	Location
fix	gene5832	fixK	bja：bll3466	CRP/FNR family transcriptional regulator	6070103-6070786
	gene6542	fixK	bja：bll2757	CRP/FNR family transcriptional regulator	6868843-6869541
	gene6539	fixL	bja：bll2760	LuxR family， sensor kinase FixL	6866141-6867658
	gene6540	fixJ	bja：bll2759	LuxR family， response regulator FixJ	6867662-6868279
	gene7359	fixA	bja：blr2038	Flavoprotein beta subunit	7803669-7804535
	gene7895	fixA	bja：blr1377	Flavoprotein beta subunit	8425108-8425857^C
	gene7566	fixX	bja：bsr1775	Ferredoxin like protein	8067228-8067524^C
	gene7567	fixC	bja：blr1774	Flavoprotein-quinone oxidoreductase	8067563-8068870^C
	gene7568	fixB	bja：blr1773	Flavoprotein alpha subunit	8068882-8069991^C
	gene7894	fixB	bja：blr1378	Flavoprotein alpha subunit	8424164-8425108^C
nod	gene5597	nodT	bsym：CIT39_22430	Multidrug efflux system	5805752-5807308
	gene7363	nodU	bja：blr2034	Carbamoyltransferase	7809897-7811504^C
	gene7367	nodU	bjp：RN69_37980	Carbamoyltransferase	7813893-7815602^C
	gene7365	nodJ	bja：blr2031	Lipooligosaccharide transport system permease	7812179-7812967^C
	gene7366	nodI	bja：blr2030	Lipooligosaccharide transport system ATP-binding protein	7812971-7813891^C
	gene7368	nodC	bja：blr2027	N-acetylglucosaminyltransferase	7816172-7817629^C
	gene7369	nodB	bja：blr2026	Chitooligosaccharide deacetylase	7817644-7818303^C
	gene7370	nodA	bja：blr2025	Nodulation protein A	7818300-7818932^C
	gene7372	nodD	bja：bll2023	LysR family transcriptional regulator	7819726-7820670
nif	gene7360	nifA	bja：blr2037	Nif-specific regulatory protein	7805053-7806801^C
	gene7569	nifW	bja：blr1771	Nitrogenase-stabilizing/protective protein	8070942-8071283^C
	gene7571	nifQ	bju：BJ6T_80490	Nitrogen fixation protein NifQ	8071938-8072714^C
	gene7572	nifH	bja：blr1769	Nitrogenase iron protein NifH	8072861-8073745
	gene7580	nifZ	bjp：RN69_39140	Nitrogen fixation protein NifZ	8078154-8078468^C
	gene7582	nifB	bju：BJ6T_80610	Nitrogen fixation protein NifB	8078954-8080537^C
	gene7583	nifT	bja：bsr1757	Nitrogen fixation protein NifT	8081201-8081425^C
	gene7594	nifX	bja：blr1747	Nitrogen fixation protein NifX	8090471-8090866^C
	gene7595	nifN	bja：blr1746	Nitrogenase molybdenum-iron protein NifN	8090863-8092272^C
	gene7596	nifE	bja：blr1745	Nitrogenase molybdenum-cofactor synthesis protein NifE	8092282-8093925^C
	gene7597	nifK	bja：blr1744	Molybdenum-iron protein beta chain	8094018-8095574^C
	gene7598	nifD	bja：blr1743	Molybdenum-iron protein alpha chain	8095640-8097142^C