Analysis of transgene expression in transgenic plants: A dissertation submitted in partial fulfilment of the requirements for the Degree of Bachelor of Horticultural Science (Honours) in Lincoln University

Abstract

Crop Research Division, DSIR has genetically modified the potato cultivar, Iwa, with increased resistance to the herbicide, chlorsulfuron. The acetohydroxy acid synthase (AHAS) gene that confers chlorsulfuron resistance to transformed plants was fused to the glucuronidase (GUS) reporter gene, expression of the whole construct being driven by the CaMV 35S promoter. In this work, the transgene expression in transgenic potatoes was analysed. The level of AHAS expression in the transgenic plants was directly ascertained by determining the level of GUS mRNA expression using slot-blot analysis. Total plant RNA was extracted and probed with the 32P-labelled 1.8 kb Pst I - EcoR I GUS fragment excised from pKIWI 101. High background reading was shown in the autoradiograph developed. A higher stringency wash was carried out to delete unspecific binding of the probe to other plant RNA species. Results from the analysis of plant mRNA using the above mentioned methods showed strong correlation to the gene copy number as well as to the field performance of the transgenic potatoes in a small herbicide trial located at Lincoln during the 1989 season. Thus, expression level of the AHAS gene was shown to influence the degree of chlorsulfuron resistance in the transgenic plants