Multiplex PCR assay for identification of commonly used disarmed Agrobacterium tumefaciens strains
نویسندگان
چکیده
The success of Agrobacterium mediated plant transformation depends to a certain extent on appropriate selection of the A. tumefaciens strain for a particular plant species. Many stages in a plant transformation procedure are prone to bacterial contamination with similar antibiotic resistance that may compromise the identity of the A. tumefaciens strain used, in turn adversely affecting success of a transformation experiment. Different primer sets were designed to exploit genetic differences among different strains of A. tumefaciens which are commonly used for plant genetic transformation, to identity confirmation as well as to distinguish them from one another. The primer sets Ach5FtsZ-F/R specific for Ach5 and C58GlyA-F/R specific for C58 were designed on chromosomal DNA while primer sets pTiBo542-F/R and nptI-F/R specific for plasmid pTiBo542 are capable to identify and distinguish these strains from one another. These primer sets when used simultaneously in multiplex PCR, produce a pattern which uniquely identifies all these strains and distinguishes them except for GV3101 and C58C1, which can further be distinguished from each other by rifampicin screening. The multiplex PCR assay and primers being reported here serve as a valuable tool in determining the identity of A. tumefaciens strains at any stage of plant transformation procedure.
منابع مشابه
Molecular identification of agrobacterium tumefaciens containing pCAMBIA 1305.2 plasmid using multiplex PCR and Gold nanoparticles multiplex probe
Conventional microbiology methods used to detect bacteria include multiple cultures and identification processes, so the results of lab work are painstaking and time-consuming. In recent years, more and more tend to use the diagnostic tests which are based on DNA; hence, DNA diagnostic biosensors have been created to perform DNA identification better. In this study, GUS and hpt genes were used ...
متن کاملMolecular identification of agrobacterium tumefaciens containing pCAMBIA 1305.2 plasmid using multiplex PCR and Gold nanoparticles multiplex probe
Conventional microbiology methods used to detect bacteria include multiple cultures and identification processes, so the results of lab work are painstaking and time-consuming. In recent years, more and more tend to use the diagnostic tests which are based on DNA; hence, DNA diagnostic biosensors have been created to perform DNA identification better. In this study, GUS and hpt genes were used ...
متن کاملAn efficient multiplex PCR assay for early detection of Agrobacterium tumefaciens in transgenic plant materials
* Correspondence: [email protected]
متن کاملColonization of Phaseolus vulgaris nodules by Agrobacterium-like strains.
Non-nodulating Agrobacterium-like strains identified among root nodule isolates of common bean were labeled with gusA, a reporter gene encoding beta-glucuronidase (GUS). Bean plants were then co-inoculated with an infective Rhizobium strain and labeled transconjugants of Agrobacterium-like strains. Blue staining of nodules showed that Agrobacterium-like strains were able to colonize these symbi...
متن کاملNew multiplex PCR methods for rapid screening of genetically modified organisms in foods
We present novel multiplex PCR methods for rapid and reliable screening of genetically modified organisms (GMOs). New designed PCR primers targeting four frequently used GMO specific sequences permitted identification of new DNA markers, in particular 141 bp fragment of cauliflower mosaic virus (CaMV) 35S promoter, 224 bp fragment of Agrobacterium tumefaciens nopaline synthase (NOS) terminator,...
متن کامل