[go: up one dir, main page]

WO2001038492A1 - Souches d'inoculation ameliorees de bradyrhizobium japonicum - Google Patents

Souches d'inoculation ameliorees de bradyrhizobium japonicum Download PDF

Info

Publication number
WO2001038492A1
WO2001038492A1 PCT/US2000/032316 US0032316W WO0138492A1 WO 2001038492 A1 WO2001038492 A1 WO 2001038492A1 US 0032316 W US0032316 W US 0032316W WO 0138492 A1 WO0138492 A1 WO 0138492A1
Authority
WO
WIPO (PCT)
Prior art keywords
seq
strain
gene
bradyrhizobium
nucleotide sequence
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US2000/032316
Other languages
English (en)
Other versions
WO2001038492A9 (fr
Inventor
Arvind A. Bhagwat
Donald L. Keister
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
University of Maryland Baltimore
University of Maryland College Park
Original Assignee
University of Maryland Baltimore
University of Maryland College Park
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by University of Maryland Baltimore, University of Maryland College Park filed Critical University of Maryland Baltimore
Priority to AU19289/01A priority Critical patent/AU1928901A/en
Publication of WO2001038492A1 publication Critical patent/WO2001038492A1/fr
Anticipated expiration legal-status Critical
Publication of WO2001038492A9 publication Critical patent/WO2001038492A9/fr
Ceased legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C12N1/205—Bacterial isolates
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales

Definitions

  • inoculants are used in several forms such as mixed with peat as a carrier, coated on seeds or directly as a liquid inoculant.
  • peat as a carrier
  • the inoculant strain typically forms only a small number of the nodules on the roots of the soybean plant (e.g. Kvien et al, 1981; Ellis et al, 1984; Howie et al, 1987).
  • the indigenous soil strains form the bulk of the nodules. This is known as the problem of competition.
  • the indigenous strains have unknown characteristics which give them an advantage in nodulation.
  • inoculant strains whether natural or genetically enhanced for superior nitrogen fixation, do not substantially enhance plant productivity since they are incapable of forming the bulk of the nodules on the plant's roots.
  • This invention addresses that limitation by having enhanced competitiveness due to mutation with transposon Tn5.
  • the present invention is directed to an isolated strain of Bradyrhizobium having increased nodulation characteristics, wherein a gene comprising a nucleotide sequence that hybridizes to the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO:2 under 5X SSC and 42°C wash conditions is not expressed.
  • the strain may include a gene that may at least partially comprise SEQ ID NO:l or SEQ ID NO:2.
  • the gene may comprise SEQ ID NO. and SEQ ID NO:2, wherein SEQ ID NO:l is located upstream of SEQ ID NO:2.
  • the present invention is also directed to a strain of Bradyrhizobium having a deposit number of NRRL-B-30052 or NRRL-B-30053.
  • the invention is directed to a method for promoting nodulation of a leguminous plant, comprising: (i) inoculating a plant or a place near a plant root with a nodulating effective amount of an inoculum comprising an isolated strain of Bradyrhizobium having increased nodulation characteristics, wherein a gene comprising a nucleotide sequence that hybridizes to the nucleotide sequence of SEQ ID NO:l or SEQ ID NO:2 under 5X SSC and 42°C wash conditions is not expressed, and
  • leguminous plant may be soybean, cowpea mungbean or siratro.
  • the invention is also directed to an isolated gene comprising a nucleotide sequence that hybridizes to the nucleotide sequence of SEQ ID NO:l or SEQ ID NO:2 under 5X SSC and 42°C wash conditions.
  • the gene may comprise at least in part SEQ ID NO:l or SEQ ID NO:2.
  • the gene also may comprise SEQ ID NO: 1 and SEQ ID NO:2, wherein SEQ ID NO:l is located upstream of SEQ ID NO:2.
  • the invention is also directed to a plant seed coated with any of the Bradyrhizobium strain discussed above.
  • the invention is directed to a composition
  • a composition comprising an isolated strain of Bradyrhizobium having increased nodulation characteristics, wherein a gene comprising a nucleotide sequence that hybridizes to the nucleotide sequence of SEQ ID NO:l or SEQ ID NO:2 under 5X SSC and 42°C wash conditions is not expressed, and an agriculturally acceptable carrier thereof.
  • Fig. 1 shows nodule occupancy of 1-110 ARS and BJ5019 when mixed together with varying proportions of USDA 438 and inoculated onto soybean plants.
  • Fig. 2 shows nodule Occupancy of BJ5019 when mixed with other B. japonicum strains at a ratio of 1 : 100 (B J5019:other strain).
  • Fig. 3 shows nodule Occupancy of BJ5019 on other host plants in competition with 1-110 ARS.
  • Fig. 4 shows the flanking regions of the Tn5 insertion site (SEQ ID NOS:l and 2).
  • Table 1 shows nodulation of soybeans by mixtures of BJ5019 and TA11 Nod +
  • Table 2 shows nodulation of soybeans by mixtures of BJ5019 and B. japonicum strain 119
  • genetic element may be any nucleotide sequence that affects the expression of a particular gene or phenotype.
  • a gene may be a genetic element.
  • a gene is a unit of nucleotide sequence that encodes a protein.
  • the gene is at least partly comprised of SEQ ID NO:l or SEQ ID NO:2, or both.
  • the gene may also comprise SEQ ID NO:l and SEQ ID NO:2, wherein SEQ ID NO:l is upstream of SEQ ID NO:2.
  • SEQ ID NO:l and SEQ ID NO:2 may be contiguous.
  • this gene is derived from Bradyhizobium.
  • the Bradyrizobium is Bradyrhizobium elkanii or Bradyrhizobium japonicum.
  • the gene is derived from B. japonicum.
  • the gene that hybridizes to SEQ ID NO:l and/or SEQ ID NO:2 may be varied, as certain non-functional variations are possible without affecting the activity of the gene.
  • the gene may contain sequences that preferably hybridize to the SEQ ID NO:l or SEQ ID NO:2 or both in either contiguous or individual form, under 5X SSC and 42°C wash conditions.
  • the genetic element may be mutated.
  • the mutation may be made so that the gene is either not expressed, which is sometimes termed knock-out mutant, or expressed in low amounts.
  • the mutation may be caused by chemical means such as EMS or MMS mutagenesis, or by sited directed mutagenesis such as by insertion of a disrupting genetic sequence. Other methods include the insertion of transposons or insertion sequences.
  • the insertion sequence may contain a sequence that codes for a selective marker, such as an antibiotic.
  • serocluster 123 strains of B. japonicum are the dominant indigenous strains and are highly competitive.
  • USDA 438 is a representative of serocluster 123.
  • the nodule occupancy of BJ5019 and 1-110 ARS when mixed with various proportions of USDA 438 is shown in Fig. 1.
  • 1-110 ARS is a derivative of USDA 110 which was selected for spontaneous resistance to azide, rifampin and streptomycin (USDA/ARS Rhizobium Germplasm Resource Collection) and is useful as a substitute for USDA 110 because it is easily distinguished due to the strain's resistance to those compounds.
  • BJ5019 The number of BJ5019 cells required for 50% nodule occupancy is more than 100-fold less than with 1-110 ARS; and for 25% nodule occupancy is almost 1000-fold less. Thus, in this type of experiment, BJ5019 is markedly enhanced for competitiveness relative to a derivative of the parent strain.
  • B. japonicum strains 119 (Patent No. 4,863,866), and TA11 Nod + (Patent No. 5,021,076) are claimed to be enhanced in nodulation and/or nitrogen fixation. Both of these strains were derived from USDA 110. Tables 1 and 2 illustrate that BJ5019 is superior to these strains in nodulation ability at a ratio of 1:1 in the inoculum; BJ5019 formed 81% of the nodules on soybean when mixed with strain 119 (U.S. Patent No. 4,863,866 herein incorporated by reference) (Table 2) and 90% of the nodules when mixed with TA11 Nod + (U.S. Patent No. 5,021,076 herein incorporated by reference)(Table 1).
  • strain TAl 1 Nod + The antibiotic resistance markers on strain TAl 1 Nod + are rifampicin, streptomycin, and azide.
  • Strain BJ5019 is resistant to kanamycin and streptomycin. Nodule occupancy by BJ5019 was obtained by scoring growth on media containing kanamycin, while occupancy by TAl 1 Nod + was determined by plating bacteroids isolated from nodules on rifampicin containing media.
  • strain 119 The antibiotic marker on strain 119 is kanamycin resistant.
  • Strain BJ5019 is resistant to kanamycin and streptomycin. Nodule occupancy by BJ5019 was obtained by scoring growth on media containing kanamycin. Independent analysis of nodule occupancy by strain 119 was not technically possible due to limitations of antibiotic marker selection and serotyping (both strains belong to the same serogroup, e.g., USDA 110).
  • B. japonicum nodulates other legume plants in addition to soybean. That the enhanced competitive nodulation phenotype is not specific to soybean is illustrated in Fig. 3.
  • BJ5019 formed more than 80% of the nodules on these legumes when mixed at a 100:1 ratio (1-110ARS: BJ5019).
  • compositions containing the Tn5 mutagenized strain of B. japonicum of the invention may be used to coat seeds. Coating is accomplished by immersing seeds in a B. japonicum-Xiansposon Tn5 enriched solution.
  • Example 1 Mutagenesis of USDA 110. Bradyrhizobium japonicum strain USDA 110 was isolated from a soybean nodule and is included in a germplasm collection, USDA-ARS National Rhizobium Germplasm Collection, Beltsville, MD 20705. It has been widely used in research and in commercial inocula.
  • Transposon-5 (Tn5).
  • Transposons are naturally occurring genetic elements carrying antibiotic resistance genes.
  • Transposon Tn5 has genes which carry resistance to kanamycin and streptomycin.
  • Tn5 mutagenesis of USDA 110 was performed by triparental mating as described by Ditta (1986). Individual colonies of USDA 110 harboring Tn5 were selected on media containing kanamycin and streptomycin (at 200 ⁇ g/ml) (Bhagwat et al, 1991).
  • Example 2 Selection of competitive nodulation phenotype. Tn5 mutants were screened for their competitive nodulation ability on the host plant Glycine max (soybean) cv. Williams. Nodulation assays were performed in sterile Leonard jar assemblies (Vincent, 1970). Soybean seeds were inoculated with a mixture of B. japonicum strains (various Tn5 mutants and strain Mc617, in the ratio of 1 : 100). The strain Mc617 is a Fix " mutant of USDA 110 (Bhagwat et al, 1991) and forms large numbers of ineffective nodules which are unable to fix N 2 .
  • Soybean plants were maintained on N-free nutrient solution for 4 weeks in a greenhouse before harvesting for nodule occupancy analysis (Bhagwat et al, 1996). Surface sterilized nodules were examined for the presence of kanamycin and streptomycin resistant bacteria to determine competitiveness of Tn5 mutant strains. The Tn5 mutant which formed the greatest number of nodules (BJ5009) was selected. This strain was deposited with the Agricultural Research Service Culture Collection (NRRL) under provisions of the Budapest treaty and assigned NRRL No. B-30052. All restrictions imposed by the depositor on the availability to the public of B-30052 will be irrevocably removed upon granting the patent.
  • NRRL Agricultural Research Service Culture Collection
  • Example 3 Confirmation that the phenotype was due to the Tn5 insertion.
  • the selected mutant strain BJ5009 was confirmed to carry a single copy of Tn5 and the vector sequences were absent.
  • the genomic DNA of BJ5009 flanking the Tn5 insertion was cloned at the EcoRI site into a suicidal vector pSUP202.
  • the resultant plasmid was introduced into USDA 110 by triparental mating (Ditta, 1986) and exconjugants resistant to kanamycin and streptomycin (Tn5 markers) but sensitive to tetracycline (pSUP202 marker) were selected.
  • the selected exconjugants were screened to confirm the Tn5 insertion into an identical EcoRl fragment as observed in BJ5009 and one such exconjugant was designated as BJ5019.
  • This strain was deposited with the Agricultural Research Service Culture Collection (NRRL) under provisions of the Budapest treaty and assigned NRRL No. B-30053. All restrictions imposed by the depositor on the availability to the public of B-30053 will be irrevocably removed upon granting the patent.
  • strain BJ5019 was indistinguishable in competitive nodulation phenotype BJ5099.
  • Example 4 Cloning of the full-length Bradyrhizobium gene that hybridizes to SEQ ID NO. l or SEQ ID NO: 2.
  • the cloning of the full-length gene for which the nucleotide sequence of SEQ ID NO:l and SEQ ID NO:2 may be a part, is accomplished using well-known molecular cloning methods, such as disclosed in for instance, Sambrook et al., which is incorporated herein by reference especially with respect to the cloning the full-length gene by hybridization to known nucleotide sequences.

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Zoology (AREA)
  • General Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Wood Science & Technology (AREA)
  • Biotechnology (AREA)
  • Genetics & Genomics (AREA)
  • Virology (AREA)
  • Microbiology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Medicinal Chemistry (AREA)
  • Biochemistry (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Dentistry (AREA)
  • Biomedical Technology (AREA)
  • Environmental Sciences (AREA)
  • Plant Pathology (AREA)
  • Pest Control & Pesticides (AREA)
  • General Engineering & Computer Science (AREA)
  • Agronomy & Crop Science (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Biophysics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Molecular Biology (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

La présente invention concerne des souches mutantes de Bradyrhizobium japonicum, BJ5009 et BJ5019, obtenues par mutagenèse par transposons de souche de B. japonicum USDA 110 connue. Ces souches sont caractérisées par une compétitivité supérieure à l'égard des nodulations chez les légumineuses, en présence de la souche parentale et d'autres souches B.japonicum.
PCT/US2000/032316 1999-11-24 2000-11-24 Souches d'inoculation ameliorees de bradyrhizobium japonicum Ceased WO2001038492A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU19289/01A AU1928901A (en) 1999-11-24 2000-11-24 Improved inoculant strains of bradyrhizobium japonicum

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US16730099P 1999-11-24 1999-11-24
US60/167,300 1999-11-24

Publications (2)

Publication Number Publication Date
WO2001038492A1 true WO2001038492A1 (fr) 2001-05-31
WO2001038492A9 WO2001038492A9 (fr) 2002-11-07

Family

ID=22606790

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2000/032316 Ceased WO2001038492A1 (fr) 1999-11-24 2000-11-24 Souches d'inoculation ameliorees de bradyrhizobium japonicum

Country Status (2)

Country Link
AU (1) AU1928901A (fr)
WO (1) WO2001038492A1 (fr)

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2233458A3 (fr) * 2001-08-13 2011-02-23 Agro.Bio Hungary Kft. Microorganismes pour le traitement du sol et un procédé de leur obtention
RU2426778C2 (ru) * 2009-03-27 2011-08-20 Общество с ограниченной ответственностью "Научно-технический центр биологических технологий в сельском хозяйстве" (ООО "НТЦ БИО") Штамм клубеньковых бактерий bradyrhizobium japonicum 206 вкпм в-9505, вирулентный к районированным сортам сои
WO2014079771A1 (fr) * 2012-11-22 2014-05-30 Basf Se Mélanges pesticides
RU2588483C2 (ru) * 2011-12-16 2016-06-27 Новозимс Биоаг А/С Штамм bradyrhizobium для улучшения роста растений (варианты), композиция, содержащая указанный штамм, и семя, покрытое композицией
RU2611199C2 (ru) * 2011-03-31 2017-02-21 Новозаймз Байолоджикалз, Инк Конкурентоспособные и эффективные штаммы бактерий

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4863866A (en) * 1987-03-12 1989-09-05 Lipha Chemicals, Inc. Bradyrhizobium japonicum mutants exhibiting superior soybean nodulation
US5021076A (en) * 1989-03-17 1991-06-04 The United States Of America As Represented By The Secretary Of Agriculture Enhancement of nitrogen fixation with Bradyrhizobium japonicum mutants

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4863866A (en) * 1987-03-12 1989-09-05 Lipha Chemicals, Inc. Bradyrhizobium japonicum mutants exhibiting superior soybean nodulation
US5021076A (en) * 1989-03-17 1991-06-04 The United States Of America As Represented By The Secretary Of Agriculture Enhancement of nitrogen fixation with Bradyrhizobium japonicum mutants

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
STACEY G.: "Bradyrhizobium japonicum nodulation genetics", FEMS MICROBIOL. LETTERS, vol. 127, 1995, pages 1 - 9, XP002939280 *

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2233458A3 (fr) * 2001-08-13 2011-02-23 Agro.Bio Hungary Kft. Microorganismes pour le traitement du sol et un procédé de leur obtention
HRP20040126B1 (hr) * 2001-08-13 2014-02-14 Agro-Bio Hungary Kft. Mikroorganizmi za tretiranje tla i postupci za njihovo dobivanje
RU2426778C2 (ru) * 2009-03-27 2011-08-20 Общество с ограниченной ответственностью "Научно-технический центр биологических технологий в сельском хозяйстве" (ООО "НТЦ БИО") Штамм клубеньковых бактерий bradyrhizobium japonicum 206 вкпм в-9505, вирулентный к районированным сортам сои
RU2611199C2 (ru) * 2011-03-31 2017-02-21 Новозаймз Байолоджикалз, Инк Конкурентоспособные и эффективные штаммы бактерий
RU2588483C2 (ru) * 2011-12-16 2016-06-27 Новозимс Биоаг А/С Штамм bradyrhizobium для улучшения роста растений (варианты), композиция, содержащая указанный штамм, и семя, покрытое композицией
WO2014079771A1 (fr) * 2012-11-22 2014-05-30 Basf Se Mélanges pesticides

Also Published As

Publication number Publication date
WO2001038492A9 (fr) 2002-11-07
AU1928901A (en) 2001-06-04

Similar Documents

Publication Publication Date Title
Stępkowski et al. European origin of Bradyrhizobium populations infecting lupins and serradella in soils of Western Australia and South Africa
Squartini et al. Rhizobium sullae sp. nov.(formerly'Rhizobium hedysari'), the root-nodule microsymbiont of Hedysarum coronarium L.
Mostasso et al. Selection of bean (Phaseolus vulgaris L.) rhizobial strains for the Brazilian Cerrados
Eardly et al. Phylogenetic position of Rhizobium sp. strain Or 191, a symbiont of both Medicago sativa and Phaseolus vulgaris, based on partial sequences of the 16S rRNA and nifH genes
Liefting et al. Sequence heterogeneity in the two 16S rRNA genes of Phormium yellow leaf phytoplasma
Sullivan et al. Four unnamed species of nonsymbiotic rhizobia isolated from the rhizosphere of Lotus corniculatus
Wei et al. Rhizobium indigoferae sp. nov. and Sinorhizobium kummerowiae sp. nov., respectively isolated from Indigofera spp. and Kummerowia stipulacea.
Anyango et al. Diversity of rhizobia nodulating Phaseolus vulgaris L. in two Kenyan soils with contrasting pHs
Dekkers et al. A two-component system plays an important role in the root-colonizing ability of Pseudomonas fluorescens strain WCS365
Brom et al. In Rhizobium etli symbiotic plasmid transfer, nodulation competitivity and cellular growth require interaction among different replicons
Devine et al. Host genetic control of symbiosis in soybean (Glycine max L.)
Villegas et al. Nitrogen-fixing sinorhizobia with Medicago laciniata constitute a novel biovar (bv. medicaginis) of S. meliloti
Gordon et al. An experimental test of the rhizopine concept in Rhizobium meliloti
Mwenda et al. Genetic diversity and symbiotic effectiveness of Phaseolus vulgaris-nodulating rhizobia in Kenya
Han et al. Molecular diversity and phylogeny of rhizobia associated with wild legumes native to Xinjiang, China
Sánchez-Cañizares et al. Endosymbiotic bacteria nodulating a new endemic lupine Lupinus mariae-josephi from alkaline soils in Eastern Spain represent a new lineage within the Bradyrhizobium genus
Neves et al. Diversity and adaptability of soybean and cowpea rhizobia in tropical soils
Belhadi et al. Vicia faba L. in the Bejaia region of Algeria is nodulated by Rhizobium leguminosarum sv. viciae, Rhizobium laguerreae and two new genospecies
Bala et al. Relationships between rhizobial diversity and host legume nodulation and nitrogen fixation in tropical ecosystems
Msaddak et al. Definition of two new symbiovars, sv. lupini and sv. mediterranense, within the genera Bradyrhizobium and Phyllobacterium efficiently nodulating Lupinus micranthus in Tunisia
León-Barrios et al. Ensifer meliloti bv. lancerottense establishes nitrogen-fixing symbiosis with Lotus endemic to the Canary Islands and shows distinctive symbiotic genotypes and host range
Strain et al. Genetic structure of Rhizobium leguminosarum biovar trifolii and viciae populations found in two Oregon soils under different plant communities
Liu et al. Nodulating competitiveness of a nonmotile Tn 7 mutant of Bradyrhizobium japonicum in nonsterile soil
Anyango et al. Competition in Kenyan soils between Rhizobium leguminosarum biovar phaseoli strain Kim5 and R. tropici strain CIAT899 using the gusA marker gene
Mahdhi et al. Characterization of root-nodulating bacteria on Retama raetam in arid Tunisian soils

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AE AG AL AM AT AU AZ BA BB BG BR BY BZ CA CH CN CR CZ DE DK DM DZ EE ES FI GB GD GE GH GM HR HU ID IL IN IS JP KE KG KR KZ LC LK LR LS LT LU LV MA MD MG MK MN MW MX MZ NO NZ PL PT RO RU SD SE SG SI SK SL TJ TM TR TT TZ UA UG US UZ VN YU ZA ZW

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): GH GM KE LS MW MZ SD SL SZ TZ UG ZW AM AZ BY KG KZ MD RU TJ TM AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE TR BF BJ CF CG CI CM GA GN GW ML MR NE SN TD TG

121 Ep: the epo has been informed by wipo that ep was designated in this application
WWE Wipo information: entry into national phase

Ref document number: 09889883

Country of ref document: US

REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

AK Designated states

Kind code of ref document: C2

Designated state(s): AE AG AL AM AT AU AZ BA BB BG BR BY BZ CA CH CN CR CZ DE DK DM DZ EE ES FI GB GD GE GH GM HR HU ID IL IN IS JP KE KG KR KZ LC LK LR LS LT LU LV MA MD MG MK MN MW MX MZ NO NZ PL PT RO RU SD SE SG SI SK SL TJ TM TR TT TZ UA UG US UZ VN YU ZA ZW

AL Designated countries for regional patents

Kind code of ref document: C2

Designated state(s): GH GM KE LS MW MZ SD SL SZ TZ UG ZW AM AZ BY KG KZ MD RU TJ TM AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE TR BF BJ CF CG CI CM GA GN GW ML MR NE SN TD TG

122 Ep: pct application non-entry in european phase