scholarly journals Assessment of cry1 Gene Contents of Bacillus thuringiensis Strains by Use of DNA Microarrays

2005 ◽  
Vol 71 (9) ◽  
pp. 5391-5398 ◽  
Author(s):  
Jaroslaw Letowski ◽  
Alejandra Bravo ◽  
Roland Brousseau ◽  
Luke Masson
Keyword(s):  
Bacillus Thuringiensis ◽  
Dna Microarrays ◽  
Specific Gene ◽  
Sequence Alignments ◽  
Fast Screening ◽  
Strain Collection ◽  
Positive Hybridization ◽  
Excellent Tool ◽  
Positive Results ◽  
Cry1 Gene

ABSTRACT A single Bacillus thuringiensis strain can harbor numerous different insecticidal crystal protein (cry) genes from 46 known classes or primary ranks. The cry1 primary rank is the best known and contains the highest number of cry genes which currently totals over 130. We have designed an oligonucleotide-based DNA microarray (cryArray) to test the feasibility of using microarrays to identify the cry gene content of B. thuringiensis strains. Specific 50-mer oligonucleotide probes representing the cry1 primary and tertiary ranks were designed based on multiple cry gene sequence alignments. To minimize false-positive results, a consentaneous approach was adopted in which multiple probes against a specific gene must unanimously produce positive hybridization signals to confirm the presence of a particular gene. In order to validate the cryArray, several well-characterized B. thuringiensis strains including isolates from a Mexican strain collection were tested. With few exceptions, our probes performed in agreement with known or PCR-validated results. In one case, hybridization of primary- but not tertiary-ranked cry1I probes indicated the presence of a novel cry1I gene. Amplification and partial sequencing of the cry1I gene in strains IB360 and IB429 revealed the presence of a cry1Ia gene variant. Since a single microarray hybridization can replace hundreds of individual PCRs, DNA microarrays should become an excellent tool for the fast screening of new B. thuringiensis isolates presenting interesting insecticidal activity.

Characterization of cry Genes in a Mexican Bacillus thuringiensis Strain Collection

1998 ◽  
Vol 64 (12) ◽  
pp. 4965-4972 ◽  
Author(s):  
Alejandra Bravo ◽  
Sergio Sarabia ◽  
Lorena Lopez ◽  
Hernesto Ontiveros ◽  
Carolina Abarca ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Specific Primers ◽  
Cry Proteins ◽  
Pcr Product ◽  
Strain Collection ◽  
Lepidopteran Insects ◽  
Ecological Patterns ◽  
Biogeographical Regions ◽  
Selection Of

ABSTRACT Mexico is located in a transition zone between the Nearctic and Neotropical biogeographical regions and contains a rich and unique biodiversity. A total of 496 Bacillus thuringiensis strains were isolated from 503 soil samples collected from the five macroregions of the country. The characterization of the strain collection provided useful information on the ecological patterns of distribution of B. thuringiensis and opportunities for the selection of strains to develop novel bioinsecticidal products. The analysis of the strains was based on multiplex PCR with novel general and specific primers that could detect the cry1,cry3, cry5, cry7, cry8,cry9, cry11, cry12,cry13, cry14, cry21, andcyt genes. The proteins belonging to the Cry1 and Cry9 groups are toxic for lepidopteran insects. The Cry3, Cry7, and Cry8 proteins are active against coleopteran insects. The Cry5, Cry12, Cry13, and Cry14 proteins are nematocidal. The Cry11, Cry21, and Cyt proteins are toxic for dipteran insects. Six pairs of general primers are used in this method. Strains for which unique PCR product profiles were obtained with the general primers were further characterized by additional PCRs with specific primers. Strains containingcry1 genes were the most abundant in our collection (49.5%). Thirty-three different cry1-type profiles were identified. B. thuringiensis strains harboringcry3 genes represented 21.5% of the strains, and 7.9% of the strains contained cry11 and cyt genes.cry7, cry8, and cry9 genes were found in 0.6, 2.4, and 2.6% of the strains, respectively. No strains carrying cry5, cry12, cry13,cry14, or cry21 genes were found. Finally, 14% of the strains did not give any PCR product and did not react with any polyclonal antisera. Our results indicate the presence of strains that may harbor potentially novel Cry proteins as well as strains with combinations of less frequently observed cry genes.

scholarly journals Bacillus thuringiensis isolates entomopathogenic for Culex quinquefasciatus (Diptera: Culicidae) and Anticarsia gemmatalis (Lepidoptera: Noctuidae)

2010 ◽  
Vol 70 (4) ◽  
pp. 1039-1046 ◽  
Author(s):  
V. Gobatto ◽  
SG. Giani ◽  
M. Camassola ◽  
AJP. Dillon ◽  
A. Specht ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Culex Quinquefasciatus ◽  
Reference Strain ◽  
Probit Analysis ◽  
Anticarsia Gemmatalis ◽  
Sds Page ◽  
Pathogenicity Tests ◽  
Lepidoptera Noctuidae ◽  
Reference Strains ◽  
Cry1 Gene

Samples of the Bacillus thuringiensis (Bt) were collected from soil and insects. Eight isolates were selected from rural soil, 15 from urban soil and 11 from insects. These were evaluated for entomopathogenicity against larvae of Anticarsia gemmatalis and Culex quinquefasciatus. The pathogenicity tests showed that a higher percentage of isolates were active against A. gemmatalis (60%) compared to C. quinquefasciatus (31%). Probit analysis (LC50) indicated that against A. gemmatalis four of the isolates presented values similar to the reference strain against A. gemmatalis, while against C. quinquefasciatus one isolate showed an LC50 similar to the reference strain (IPS-82). SDS-PAGE characterisation of two isolates showed a 27 kDa protein fraction related to the Bt subspecies israelensis cytolytic toxin (cyt) gene. One 130 kDa protein, possibly related to the Bt crystal inclusions (cry1) gene, was identified in the other two isolates, which were more toxic for lepidoptera; another isolate presented a protein of 100 kDa. Some new local Bt isolates had similar LC50 probit values to the reference strains.

An historical and genomic view of schistosome conjugal biology with emphasis on sex-specific gene expression

Parasitology ◽  
2004 ◽  
Vol 128 (S1) ◽  
pp. S11-S22 ◽  
Author(s):  
K. F. HOFFMANN
Keyword(s):  
Dna Microarrays ◽  
Historical Review ◽  
Transcriptional Gene Silencing ◽  
Specific Gene ◽  
Post Transcriptional Gene Silencing ◽  
Genomic Tools ◽  
Tools And Techniques ◽  
Sexually Mature ◽  
Identification And Characterization

The genetic programmes associated with the sexual biology of dioecious schistosomes remain a critically important but significantly understudied area of parasitology. Throughout the last four decades, progress has been slow in describing the gross antigenic and proteomic differences linked to sexually mature schistosomes and in characterizing some of the sex-associated transcripts and regulatory mechanisms induced during developmental maturation. These investigations have been severely hindered by the lack of complete EST/genomic information, as well as corresponding post- and functional-genomic tools for studying these pathogenic parasites. As near complete transcriptomes forSchistosoma japonicumandS. mansonihave recently been reported, and both DNA microarrays and post-transcriptional gene silencing have been applied to schistosomes, the tools and techniques for the high-throughput identification and characterization of transcripts involved in conjugal biology are now readily available. Here, an historical review is presented that summarizes some of the most significant findings associated with schistosome sex and sexual maturation during the last several decades. Following this discussion is a current overview of some modern day genomic approaches used to study schistosomes, which illustrates how major advances in the field of conjugal biology will be achieved.

scholarly journals Amplifikasi Gen CRY1 dan Analisis Genom Isolat Bacillus thuringiensis Lokal

2009 ◽  
Vol 15 (1) ◽  
pp. 1-4
Author(s):  
Dwi Suryanto
Keyword(s):  
Bacillus Thuringiensis ◽  
Plutella Xylostella ◽  
Genome Analysis ◽  
Gel Electrophoresis ◽  
Heliothis Armigera ◽  
Pulsed Field Gel Electrophoresis ◽  
Commercial Strain ◽  
Genome Profiles ◽  
Cry1 Gene ◽  
Culex Sp

A study on amplification of Cry1 gene and genome analysis of local isolate of Bacillus thuringiensis TU1 has been done. Amplification was done for cryI gene by PCR-technique. Genome analysis was performed using pulsed-field gel electrophoresis. Insecticidal specificity assay of the isolate to larvae was done in larvae of Heliothis armigera, Plutella xylostella, Aedes aegypti, and Culex sp. Isolate of commercial strain, B. thuringiensis var kurstaki strain HD-7, was used as control for amplification of cryI gene, genome analysis, and insecticidal specificity assay. The result showed that insecticidal specificity assay of the isolates of both TU1 and commercial have similar spectrum of toxicity to the larvae. Amplification of cry1 gene resulted to a fragment of approximately 550 bp in both TU1 and commercial. Genome profiles of TU1 and commercial strain were similar.

scholarly journals Low prevalence of Moraxella catarrhalis in the patients who suffered from conjunctivitis in the southwest of Iran

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Ahmad Farajzadeh Sheikh ◽  
Mustafa Feghhi ◽  
Maryam Torabipour ◽  
Morteza Saki ◽  
Hojat Veisi
Keyword(s):  
Moraxella Catarrhalis ◽  
Positive Culture ◽  
Culture Method ◽  
Specific Gene ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Southwest Of Iran ◽  
Human Infections ◽  
Low Prevalence ◽  
Positive Results

Abstract Objective Moraxella catarrhalis is a non-motile Gram-negative diplococcus bacterium that contributed to several human infections including conjunctivitis. This study aimed to reveal the prevalence of M. catarrhalis in patients who suffered from conjunctivitis in Ahvaz city, southwest of Iran. Results Out of 100 conjunctiva swab specimens, M. catarrhalis was isolated only from one (1%) conjunctivitis cases using the culture method. This strain was isolated from a 34 years old female patient. Also, the results of the polymerase chain reaction (PCR) were in agreement with the culture method, and the specimen that showed positive culture was also positive for specific gene of M. catarrhalis. The remaining 99 specimens did not show positive results with any of the culture and PCR methods.

scholarly journals Novel Cell Wall Hydrolase CwlC fromBacillus thuringiensisIs Essential for Mother Cell Lysis

2018 ◽  
Vol 84 (7) ◽  
Author(s):  
Xiaomin Chen ◽  
Tantan Gao ◽  
Qi Peng ◽  
Jie Zhang ◽  
Yunrong Chai ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bacillus Thuringiensis ◽  
Insecticidal Activity ◽  
Mother Cell ◽  
Uv Light ◽  
Point Mutations ◽  
Cell Lysis ◽  
Specific Gene ◽  
Content Type ◽  
Cell Wall Hydrolase

ABSTRACTIn this study, a sporulation-specific gene (tentatively namedcwlC) involved in mother cell lysis inBacillus thuringiensiswas characterized. The encoded CwlC protein consists of an N-terminalN-acetylmuramoyl-l-alanine amidase (MurNAc-LAA) domain and a C-terminal amidase02 domain. The recombinant histidine-tagged CwlC proteins purified fromEscherichia coliwere able to directly bind to and digest theB. thuringiensiscell wall. The CwlC point mutations at the two conserved glutamic acid residues (Glu-24 and Glu-140) shown to be critical for the catalytic activity in homologous amidases resulted in a complete loss of cell wall lytic activity, suggesting that CwlC is anN-acetylmuramoyl-l-alanine amidase. Results of transcriptional analyses indicated thatcwlCis transcribed as a monocistronic unit and that its expression is dependent on sporulation sigma factor K (σK). Deletion ofcwlCcompletely blocked mother cell lysis during sporulation without impacting the sporulation frequency, Cry1Ac protein production, and insecticidal activity. Taken together, our data suggest that CwlC is an essential cell wall hydrolase forB. thuringiensismother cell lysis during sporulation. EngineeredB. thuringiensisstrains targetingcwlC, which allows the crystal inclusion to remain encapsulated in the mother cell at the end of sporulation, may have the potential to become more effective biological control agents in agricultural applications since the crystal inclusion remains encapsulated in the mother cell at the end of sporulation.IMPORTANCEMother cell lysis has been well studied inBacillus subtilis, which involves three distinct yet functionally complementary cell wall hydrolases. In this study, a novel cell wall hydrolase, CwlC, was investigated and found to be essential for mother cell lysis inBacillus thuringiensis. CwlC ofB. thuringiensisonly shows 9 and 21% sequence identity with knownB. subtilismother cell hydrolases CwlB and CwlC, respectively, suggesting that mechanisms of mother cell lysis may differ betweenB. subtilisandB. thuringiensis. ThecwlCgene deletion completely blocked the release of spores and crystals from the mother cell without affecting insecticidal activity. This may provide a new effective strategy for crystal encapsulation against UV light inactivation.

scholarly journals The Bacillus thuringiensis cyt Genes for Hemolytic Endotoxins Constitute a Gene Family

2001 ◽  
Vol 67 (3) ◽  
pp. 1090-1096 ◽  
Author(s):  
Alejandra Guerchicoff ◽  
Armelle Delécluse ◽  
Clara P. Rubinstein
Keyword(s):  
Bacillus Thuringiensis ◽  
Gene Family ◽  
Dna Sequences ◽  
Biologically Active ◽  
Sequence Alignments ◽  
Structural Domains ◽  
Toxic Activity ◽  
Pcr Screening ◽  
High Degree

ABSTRACT In the same way that cry genes, coding for larvicidal delta endotoxins, constitute a large and diverse gene family, thecyt genes for hemolytic toxins seem to compose another set of highly related genes in Bacillus thuringiensis. Although the occurrence of Cyt hemolytic factors in B. thuringiensishas been typically associated with mosquitocidal strains, we have recently shown that cyt genes are also present in strains with different pathotypes; this is the case for themorrisoni subspecies, which includes strains biologically active against dipteran, lepidopteran, and coleopteran larvae. In addition, while one Cyt type of protein has been described in all of the mosquitocidal strains studied so far, the present study confirms that at least two Cyt toxins coexist in the more toxic antidipteran strains, such as B. thuringiensis subsp.israelensis and subsp. morrisoni PG14, and that this could also be the case for many others. In fact, PCR screening and Western blot analysis of 50 B. thuringiensis strains revealed that cyt2-related genes are present in all strains with known antidipteran activity, as well as in some others with different or unknown host ranges. Partial DNA sequences for several of these genes were determined, and protein sequence alignments revealed a high degree of conservation of the structural domains. These findings point to an important biological role for Cyt toxins in the final in vivo toxic activity of many B. thuringiensis strains.

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