High-Throughput and Accurate Determination of Transgene Copy Number and Zygosity in Transgenic Maize: From DNA Extraction to Data Analysis.

ABSTRACT: It is vital to develop high-throughput methods to determine transgene copy numbers initially and zygosity during subsequent breeding. In this study, the target sequence of the previously reported endogenous reference gene hmg was analyzed using 633 maize inbred lines, and two SNPs were observed. These SNPs significantly increased the PCR efficiency, while the newly developed hmg gene assay (hmg-taq-F2/R2) excluding these SNPs reduced the efficiency into normal ranges. The TaqMan amplification efficiency of bar and hmg with newly developed primers was calculated as 0.993 and 1.000, respectively. The inter-assay coefficient of variation (CV) values for the bar and hmg genes varied from 1.18 to 2.94%. The copy numbers of the transgene bar using

SUBMITTER: Liu F 

PROVIDER: S-EPMC8619409 | biostudies-literature | 2021 Nov

REPOSITORIES: biostudies-literature