A High-Throughput Standard PCR-Based Genotyping Method for Determining Transgene Zygosity in Segregating Plant Populations.

ABSTRACT: In crop research programs that implement transgene-based strategies for trait improvement it is necessary to distinguish between transgene homozygous and hemizygous individuals in segregating populations. Direct methods for determining transgene zygosity are technically challenging, expensive, and require specialized equipment. In this report, we describe a standard PCR-based protocol coupled with capillary electrophoresis that can identify transgene homozygous and hemizygous individuals in a segregating population without knowledge of transgene insertion site. PCR primers were designed to amplify conserved T-DNA segments of the 35S promoter, OCS terminator, and NPTII kanamycin resistance gene in the pHellsgate-8 RNAi construct for the Gossypium hirsutum phytochrome A1 gene. Using a

SUBMITTER: Geng L 

PROVIDER: S-EPMC5522864 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature