Activity-Based Gene Cloning through PCR Module as a Part of Undergraduate Minor Project in Biotechnology

Abstract

Identification and characterization of native isolates from environmental sources assumes immense significance from research, and commercial perspective. Microbial identification using a molecular biology approach based on 16S rDNA has been the popular and well-established method. Experiments of Molecular Biology and rDNA Technology have always been a challenge for effective teaching owing to the cost, time, inconsistent results and expertise. The study was undertaken with a broad objective of giving a hands-on experience of selected molecular biology and rDNA technology experiments and related activities to the undergraduate students of Biotechnology Engineering, as a module of Minor project identified as flagship course. The genomic DNA of selected environmental soil isolates was isolated by the conventional lysis method, followed by amplification of its 16S rDNA by PCR, using universal primers. The resulting amplicons were purified, cloned into the TA vector, and analyzed for insert and the recombinant vector was transformed into host cells. The ligated cloned vector showed a size of 4500 bp as against the control vector of 3000 bp. The growth of transformants was confirmed by selective growth against ampicillin antibiotic and by blue-white screening. The exercise helped in addressing eight graduate attributes related to (investigation of technical issues, modern engineering tools, discipline-specific tools, team-work and produce technical report). Formal feedback from the students evidenced that the students had good experiential learning from the exercise. Thus, the study related to cloning exercise was instrumental in providing hands-on experience and enhanced the skill sets of the students related to fundamental molecular biology.