Developing Transgenic Plants: Through Application of Reverse Genetic Technologies Annotated edition

Plant cell wall structural proteins (CWPs) contribute to the basal defense against pathogens. Therefore, the prospect of developing disease resistant crop cultivars through genetic manipulation of CWPs is promising. Research presented here utilized a novel CW annotated Arabidopsis gene, the PELPK1 for functional characterization by using bioinformatics and computational analyses, and by applying various reverse genetics techniques, e.g., expression, mutational, and proteomic analyses. Specific techniques used are: (i) construction of several plant transformation vectors such as PELPK1-promoter:: GUS reporter fusion, PELPK1-CDS:: GFP translational fusion, RNA interference of PELPK1 + PELPK2, and constitutive over-expression of PELPK1, and development of transgenic lines harbouring the above vector constructs as well as sequence-indexed T-DNA insertions in PELPK1 gene, (ii) performing of various molecular techniques such as qRT-PCR, GUS staining, confocal laser-scanning microscopy, immunolabeling, TEM, 2D gel, LCMS/MS, and microtomy, and (iii) carrying out of genotypic and phenotypic analyses of the loss of function and gain of function mutant lines under various stress conditions