Screening of vip1/vip2 binary toxin gene and its isolation and cloning from local Bacillus thuringiensis isolates

SUMMARYEntomopathogenic bacteria produce insecticidal proteins that accumulate in inclusion bodies or parasporal crystals (such as the Cry and Cyt proteins) as well as insecticidal proteins that are secreted into the culture medium. Among the latter are the Vip proteins, which are divided into four families according to their amino acid identity. The Vip1 and Vip2 proteins act as binary toxins and are toxic to some members of the Coleoptera and Hemiptera. The Vip1 component is thought to bind to receptors in the membrane of the insect midgut, and the Vip2 component enters the cell, where it displays its ADP-ribosyltransferase activity against actin, preventing microfilament formation. Vip3 has no sequence similarity to Vip1 or Vip2 and is toxic to a wide variety of members of the Lepidoptera. Its mode of action has been shown to resemble that of the Cry proteins in terms of proteolytic activation, binding to the midgut epithelial membrane, and pore formation, although Vip3A proteins do not share binding sites with Cry proteins. The latter property makes them good candidates to be combined with Cry proteins in transgenic plants (Bacillus thuringiensis-treated crops [Bt crops]) to prevent or delay insect resistance and to broaden the insecticidal spectrum. There are commercially grown varieties of Bt cotton and Bt maize that express the Vip3Aa protein in combination with Cry proteins. For the most recently reported Vip4 family, no target insects have been found yet.

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“…So far, toxicity against 10 coleopteran species (7,11,12,24,37,38) and the hemipteran species Aphis gossypii (13,14) has been found.…”

Section: Insecticidal Activitymentioning

confidence: 99%

Bacterial Vegetative Insecticidal Proteins (Vip) from Entomopathogenic Bacteria

Chakroun

Banyuls

Bel

et al. 2016

Microbiol Mol Biol Rev

243

261

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“…Agrobacterium -mediated transformation in safflower was limited only to selectable marker and reporter gene constructs (Ying et al 1992 ; Rohini and Rao 2000 ; Belide et al 2011 ; Motamedi et al 2011 ; Sujatha et al 2012 ). Insect-resistant transgenic crops have been produced using overexpression of Bt ( Bacillus thuringiensis ) toxins, but these toxins exhibit little toxicity against such homopteran insects as aphids, mealybugs, and whiteflies (Chougule and Bonning 2012 ; Shingote et al 2013 ). We, therefore, opted for the more effective Lentil - lectin gene construct for transformation of safflower.…”

Section: Discussionmentioning

confidence: 99%

Parameters influencing Agrobacterium-mediated transformation system in safflower genotypes AKS-207 and PKV Pink

et al. 2016

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Shoot regeneration in safflower (Carthamus tinctorius ‘AKS 207’ and ‘PKV Pink’) genetically transformed using Agrobacterium was used for assessing various constraints to the efficiency of transformation including infection period, virulence induction medium, co-cultivation period, bacterial titre, selection regime, and the natural phenolic compound acetosyringone. Transformation frequency was promising with 8–10-day-old cotyledonary leaf explants. Therefore, explants of that age cultured on Agrobacterium minimal medium (AB) containing 100 µM acetosyringone were infected with Agrobacterium (cell titre 0.5 OD600nm) for 15 min followed by 48 h of co-cultivation on kanamycin-enriched medium (50 mg/L). Transformation of the shoots was confirmed using β-glucuronidase (GUS) histochemical assay and polymerase chain reaction (PCR). With the transformation protocol thus optimized, the transformation frequency as determined using GUS assays was 54.0 % for AKS 207 and 47.6 % for PKV Pink. The corresponding figures using PCR were 27.0 and 33.3 %. The transformed shoots required 10–14 weeks of culture initiation but produced very few roots.Electronic supplementary materialThe online version of this article (doi:10.1007/s13205-016-0497-4) contains supplementary material, which is available to authorized users.

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“…The binary toxins Vip2/Vip1 have been found to have specificity to Coleoptera [Arnaut et al, 2011;Bi et al, 2015;Feitelson et al, 2003;Schnepf et al, 2003;Shingote et al, 2013;Warren, 1997]. These families of genes have been observed to occur together [Hernández-Rodríguez et al, 2009].…”

Section: Pathogenicity Islandmentioning

confidence: 99%

Complete Sequence and Organization of pFR260, the <b><i>Bacillus thuringiensis</i></b> INTA Fr7-4 Plasmid Harboring Insecticidal Genes

Navas

Amadío²,

Ortiz³

et al. 2017

Microb Physiol

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We report the complete sequence and analysis of pFR260, a novel megaplasmid of 260,595 bp from the Bacillus thuringiensis strain INTA Fr7-4 isolated in Argentina. It carries 7 insecticidal genes: 3 cry8 copies previously reported, 2 vip1, and 2 vip2. Also, it carries a gene encoding a putative atypical Cry protein. These genes are arranged in a region of approximately 105 kbp in size with characteristics of a pathogenicity island with a potential coleopteran-specific insecticide profile. DNA strand composition asymmetry, as determined by GC skew analysis, and the presence of a Rep protein involved in the initiation of replication suggest a bidirectional theta mechanism of replication. In addition, many genes involved in conjugation and a CRISPR-Cas system were detected. The pFR260 sequence was deposited in GenBank under accession number KX258624.

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Screening of vip1/vip2 binary toxin gene and its isolation and cloning from local Bacillus thuringiensis isolates

Cited by 14 publications

References 18 publications

Bacterial Vegetative Insecticidal Proteins (Vip) from Entomopathogenic Bacteria

Bacterial Vegetative Insecticidal Proteins (Vip) from Entomopathogenic Bacteria

Parameters influencing Agrobacterium-mediated transformation system in safflower genotypes AKS-207 and PKV Pink

Complete Sequence and Organization of pFR260, the <b><i>Bacillus thuringiensis</i></b> INTA Fr7-4 Plasmid Harboring Insecticidal Genes

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