Open AccessA Highly Efficient Transposon Mutagenesis System for the Tomato Pathogen Clavibacter michiganensis subsp. michiganensis
Author(s) -
Oliver Kirchner,
Karl-Heinz Gartemann,
Eva-Maria Zellermann,
Rudolf Eichenlaub,
A. Burger
Publication year - 2001
Publication title -
molecular plant-microbe interactions
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.565
H-Index - 153
eISSN - 1943-7706
pISSN - 0894-0282
DOI - 10.1094/mpmi.2001.14.11.1312
Subject(s) - transposon mutagenesis , transposable element , clavibacter michiganensis , biology , transposition (logic) , mutagenesis , dna , mutant , genetics , microbiology and biotechnology , pathogen , gene , linguistics , philosophy
A transposon mutagenesis system for Clavibacter michiganensis subsp. michiganensis was developed based on antibiotic resistance transposons that were derived from the insertion element IS1409 from Arthrobacter sp. strain TM1 NCIB12013. As a prerequisite, the electroporation efficiency was optimized by using unmethylated DNA and treatment of the cells with glycine such that about 5 × 10 6 transformants per μg of DNA were generally obtained. Electroporation of C. michiganensis subsp. michiganensis with a suicide vector carrying transposon Tn1409C resulted in approximately 1 × 10 3 transposon mutants per μg of DNA and thus is suitable for saturation mutagenesis. Analysis of Tn1409C insertion sites suggests a random mode of transposition. Transposition of Tn1409C was also demonstrated for other subspecies of C. michiganensis.