Next-generation genome sequencing can be used to rapidly characterise sequences flanking T-DNA insertions in random insertional mutants of Leptosphaeria maculans.

ABSTRACT:

Background

Banks of mutants with random insertions of T-DNA from Agrobacterium tumefaciens are often used in forward genetics approaches to identify phenotypes of interest. Upon identification of mutants of interest, the flanking sequences of the inserted T-DNA must be identified so that the mutated gene can be characterised. However, for many fungi, this task is not trivial as widely used PCR-based methods such as thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR) are not successful.

Findings

Next-generation Illumina sequencing was used to locate T-DNA insertion sites in four mutants of Leptosphaeria maculans, a fungal plant pathogen. Sequence reads of up to 150 bp and coverage ranging from 6 to 24 times, were sufficient for identification of i

SUBMITTER: Chambers K 

PROVIDER: S-EPMC5611616 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature