Authors:

Connor Murray

Date Created:

Not specified

Course Title:

Professor:

Not specified

About Paper:

Species of Agrobacterium infect plant cells, transporting bacterial Transfer-DNA (T-DNA) which may then integrate into the plant chromosome. Integrated T-DNA and the disrupted plant DNA sequences often contain deletions or insertions at junctions where T-DNA is inserted into plant DNA, as well as major plant genome rearrangements such as translocations or inversions. These integration-induced changes are similar to changes in the genome that occur after repair of DNA breaks, prompting research that investigates the role of DNA repair mechanisms in T-DNA integration. This paper describes an experiment designed to test if the proteins DNA Polymerase θ (PolQ) and VirD2 interact in plants. PolQ is a low fidelity DNA polymerase that is necessary for microhomology-mediated end-joining, a highly mutagenic pathway of double strand DNA break repair. VirD2 is an Agrobacterium virulence protein that plays a key role in processing and secretion of T-DNA from Agrobacterium, and in T-DNA transport to the plant cell nucleus. Bimolecular fluorescence complementation will be used to determine if these proteins interact. In the T-DNA region of a binary vector, there is a red fluorescent nuclear marker, an nVenus-VirD2 expression cassette, and a cCFP-PolQ expression cassette that will be introduced into Agrobacterium. The resulting strain will be used to infect plants. The infection site will be analyzed by fluorescence microscopy. Infected cells will have a red fluorescent nucleus. If PolQ and VirD2 interact in the proper orientation, the attached fragments of the fluorescent protein Venus will fold and create visible yellow fluorescence.

Source:

Purdue University / 2023

Topics:

No topics listed

Co-authors:

Connor Murray