Lab Report for AGRO/BIOTC460
LAB REPORT 5
NAME: M. Azrul Akmal Saharudin
SUBMISSION DATE: May 3rd , 2011
Transgene Detection by Phenotypic Screening and Molecular Diagnosis Test.
Abstract
The introduction of Bar gene into maize and using phenotypic screening and molecular diagnosis test to detect the expression of the gene are one of commonly use method for transgene detection. We were using maize plant that was transformed with Bar gene using gene gun. For phenotypic screening, BASTA(Liberty) herbicide was applied to the plant to detect whether the plant show the phenotype of BASTA resistant or not. For molecular diagnosis test, we have used polymerase chain reaction(PCR) and gel electrophoresis to detect whether the gene present in the plant genome or not. Transgenic plants were selected and regenerated using herbicide resistance and GUS as a marker. Based on phenotypic and molecular analyses of groups putative transgenic plants confirmed the co-integration bar genes with a frequency of 54% of the transgenic plants. Another 46% carried a false positive result by showing resistant in phenotypic screening but confirmed not expressing the Bar gene by molecular diagnosis test. This result proved that transformation using gene gun produce an approximately ’50-50 chance’ for positive result because the intracellular target is random.
Introduction
Plant transformation is the introduction of a foreign gene of DNA into the host plant tissue and this foreign gene presents a specific trait1. When we talk about transformation, actually we are talking about genetic alteration of a cell resulting from the direct uptake, incorporation and expression of exogenous genetic materials from its surrounding and taken up through cell membranes. Principally, foreign gene is known as the transgene1. Plants are transformed when we want to create transgenic plant which contains gene that confers desirable traits. This is possible due to the outcome of functional...