Home LiteratureArticle Details
PMID: 3037278 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Transposon-induced symbiotic mutants of Bradyrhizobium japonicum: isolation of two gene regions essential for nodulation.

Molecular & general genetics : MGG ·Vol. 207 ·No. 1 ·1987-04-00 ·Pages 15-23

So JS, Hodgson AL, Haugland R, Leavitt M, Banfalvi Z, Nieuwkoop AJ, Stacey G

Abstract

Two strains of the soybean endosymbiont Bradyrhizobium japonicum, USDA 110 and 61 A101 C, were mutagenized with transposon Tn5. After plant infection tests of a total of 6,926 kanamycin and streptomycin resistant transconjugants, 25 mutants were identified that are defective in nodule formation (Nod-) or nitrogen fixation (Fix-). Seven Nod- mutants were isolated from strain USDA110 and from strain 61 A101 C, 4 Nod- mutants and 14 Fix- mutants were identified. Subsequent auxotrophic tests on these symbiotically defective mutants identified 4 His- Nod- mutants of USDA110. Genomic Southern analysis of the 25 mutants revealed that each of them carried a single copy of Tn5 integrated in the genome. Three 61 A101 C Fix- mutants were found to have vector DNA co-integrated along with Tn5 in the genome. Two independent DNA regions flanking Tn5 were cloned from the three non-auxotrophic Nod- mutants and one His-Nod- mutant of USDA110. Homogenotization of the cloned fragments into wild-type strain USDA110 and subsequent nodulation assay of the resulting homogenotes confirmed that the Tn5 insertion was responsible for the Nod- phenotype. Partial EcoR1 restriction enzyme maps around the Tn5 insertion sites were generated. Hybridization of these cloned regions to the previously cloned nod regions of R. meliloti and nif and nod regions of B. japonicum USDA110 showed no homology, suggesting that these regions represent new symbiotic clusters of B. japonicum.

MeSH Terms
DNA Transposable Elements DNA, Bacterial/genetics DNA, Recombinant Genes, Bacterial Nitrogen Fixation Rhizobiaceae/genetics,physiology Rhizobium/genetics Sequence Homology, Nucleic Acid Soybeans/microbiology Species Specificity Symbiosis
Chemicals
DNA Transposable Elements DNA, Bacterial DNA, Recombinant
Authors & Affiliations
7 authors, click to expand affiliations / ORCID
So J S
Hodgson A L
Haugland R
Leavitt M
Banfalvi Z
Nieuwkoop A J
Stacey G
References (31)
31 references, click to expand
  1. DNA sequence of the Rhizobium leguminosarum nodulation genes nodAB and C required for root hair curling.
    Nucleic Acids Res. 1984 Dec 21;12(24):9497-508 PMID: 6514582
  2. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I.
    J Mol Biol. 1977 Jun 15;113(1):237-51 PMID: 881736
  3. Insertion and deletion mutations within the nif region of Rhizobium japonicum.
    Plant Mol Biol. 1984 May;3(3):159-68 PMID: 24310349
  4. Broad host range DNA cloning system for gram-negative bacteria: construction of a gene bank of Rhizobium meliloti.
    Proc Natl Acad Sci U S A. 1980 Dec;77(12):7347-51 PMID: 7012838
  5. Tn5 carries a streptomycin resistance determinant downstream from the kanamycin resistance gene.
    Mol Gen Genet. 1983;191(2):288-94 PMID: 6312272
  6. Symbiotic nitrogen fixation: molecular cloning of Rhizobium genes involved in exopolysaccharide synthesis and effective nodulation.
    J Mol Appl Genet. 1982;1(6):585-96 PMID: 6296257
  7. Transposon Tn5 specifies streptomycin resistance in Rhizobium spp.
    J Bacteriol. 1984 May;158(2):580-9 PMID: 6327612
  8. Transposon Tn5-induced mutagenesis of Rhizobium japonicum yielding a wide variety of mutants.
    J Bacteriol. 1984 Jul;159(1):335-40 PMID: 6330038
  9. Molecular cloning of Rhizobium trifolii genes involved in symbiotic nitrogen fixation.
    J Mol Appl Genet. 1982;1(4):315-26 PMID: 6286830
  10. ISRm1: A Rhizobium meliloti insertion sequence that transposes preferentially into nitrogen fixation genes.
    J Mol Appl Genet. 1982;1(5):405-18 PMID: 6296251
  11. In vitro packaging of lambda and cosmid DNA.
    Methods Enzymol. 1979;68:299-309 PMID: 161604
  12. Transposon mutagenesis in rhizobia which can nodulate both legumes and the nonlegume parasponia.
    Appl Environ Microbiol. 1982 Jan;43(1):233-6 PMID: 16345924
  13. Isolation and characterization of the DNA region encoding nodulation functions in Bradyrhizobium japonicum.
    J Bacteriol. 1985 Dec;164(3):1301-8 PMID: 2999080
  14. Ineffective and non-nodulating mutant strains of Rhizobium japonicum.
    J Bacteriol. 1976 Aug;127(2):763-9 PMID: 986388
  15. Location of nodulation and nitrogen fixation genes on a high molecular weight plasmid of R. meliloti.
    Mol Gen Genet. 1981;184(2):318-25 PMID: 6949000
  16. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity.
    Anal Biochem. 1983 Jul 1;132(1):6-13 PMID: 6312838
  17. Isolation and characterization of transposon Tn5-induced symbiotic mutants of Rhizobium loti.
    J Bacteriol. 1985 Apr;162(1):335-43 PMID: 2984178
  18. Heme biosynthesis in Rhizobium. Identification of a cloned gene coding for delta-aminolevulinic acid synthetase from Rhizobium meliloti.
    J Biol Chem. 1982 Aug 10;257(15):8724-30 PMID: 7096330
  19. Detection of specific sequences among DNA fragments separated by gel electrophoresis.
    J Mol Biol. 1975 Nov 5;98(3):503-17 PMID: 1195397
  20. Physical and genetic map of a Rhizobium meliloti nodulation gene region and nucleotide sequence of nodC.
    J Bacteriol. 1985 May;162(2):469-76 PMID: 2985535
  21. DNA sequencing with chain-terminating inhibitors.
    Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463-7 PMID: 271968
  22. A restriction enzyme cleavage map of Tn5 and location of a region encoding neomycin resistance.
    Mol Gen Genet. 1979;177(1):65-72 PMID: 231729
  23. Physical organization of the Bradyrhizobium japonicum nitrogenase gene region.
    J Bacteriol. 1984 Sep;159(3):857-62 PMID: 6090394
  24. Vector insertion mutagenesis of Rhizobium sp. strain ORS571: direct cloning of mutagenized DNA sequences.
    J Bacteriol. 1985 Apr;162(1):317-23 PMID: 2984177
  25. Rhizobium japonicum derivatives differing in nitrogen-fixing efficiency and carbohydrate utilization.
    Appl Environ Microbiol. 1976 Oct;32(4):511-9 PMID: 988784
  26. Relation between Glutamine Synthetase and Nitrogenase Activities in the Symbiotic Association between Rhizobium japonicum and Glycine max.
    Plant Physiol. 1976 Apr;57(4):542-6 PMID: 16659522
  27. Host-specific nodulation is encoded on a 14kb DNA fragment in Rhizobium trifolii.
    Plant Mol Biol. 1984 Jan;3(1):3-11 PMID: 24310254
  28. Physical and genetic characterization of symbiotic and auxotrophic mutants of Rhizobium meliloti induced by transposon Tn5 mutagenesis.
    J Bacteriol. 1982 Jan;149(1):114-22 PMID: 6274841
  29. A general method for site-directed mutagenesis in prokaryotes.
    Nature. 1981 Jan 1;289(5793):85-8 PMID: 6256652
  30. Repeated sequences similar to insertion elements clustered around the nif region of the Rhizobium japonicum genome.
    J Bacteriol. 1985 May;162(2):535-42 PMID: 2985537
  31. Localization of symbiotic mutations in Rhizobium meliloti.
    J Bacteriol. 1983 Feb;153(2):635-43 PMID: 6296048
Article Info
Journal
Molecular & general genetics : MGG
Abbr.
Mol Gen Genet
ISSN
0026-8925
DOI
10.1007/BF00331485
Published
1987-04-00
Pages
15-23
Language
English
Region
Germany
NLM ID
0125036
Subset
IM
Grants
NIGMS NIH HHS · 1-RO1-GM 33494-O1A1 · United States
External Links
PubMed source DOI: 10.1007/BF00331485
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2