Modification of Ti plasmid which extracted from Agrobacteriumtumefaciens to construction the dTi vector for gene cloning

Abstract EN

A total of 47 of infected potatoes in different size (Solanumtuberosum) were collected from local vegetable and fruit market and from University of Basrah-Collage of Agriculture, Department of Plant Protection in addition to the three soil samples were taken from Basrah city.

From which 22 (44 %) isolates were isolation and identification including 19 (86.4 %) isolates from infected potatoes and 3 (13.7 %) isolates from soil.

All isolates were identification and characterization by using biochemical, physiological and biotechinqicalinvestigation.

The extracted DNA of Agrobacteriumtumefaciens isolates were subjected to PCR for amplifying 16S rRNA and for amplifying T-DNA fragment then subjected to gel electrophoresis.

The individual band of the 16S rRNA gene was characterized by 1479 bp and of the T-DNA fragment by 1200 bp.

The products were comparison with the standard molecular DNA ladder (1200 and 1500 bp).

The purified β-lactamase gene was cut from PGLO plasmid and ligated by use T4DNA ligase enzyme with the Ti plasmid which disarmed T-DNA by the same restriction enzyme.

The result was indicated by using the E.coli K12 for carries the Ti plasmid vector which contain a β-Lactamase gene when put the antibiotic ampicillin in different consternation into the LB, MHA plate only the colonies which that have picked up exogenous DNA (dTi plasmid) can grow that is mean it become resistance to ampicillin by using dTi vector.