Asian Journal of Microbiology, Biotechnology & Environmental Sciences Paper

Vol 205, Issue 4, 2018; Page No.(1286-1293)

CLONING AND EXPRESSION OF A POTENTIAL REPORTER GENE DSRED2 FOR USE IN EUKARYOTIC SYSTEM AS A DUAL-LABELING SYSTEM +

ASIT B. MANDAL, GLENN B. COLLINS, KANTI MEENA AND SOURAV DUTTA

Abstract

Transgene/s are integrated from heterologous sources into recipient cells, making the foundation of transgenic development. However, introgression of the transgene/s into the recipient system generally cannot be visualized sensu lato, which pose serious hindrance in confirming both transient/stable integration and expression of the transgene/s. Indispensability of reporter gene co-ligated in the expression vector become inevitable as integration and expression of the reporter gene is detectable visually after proper chemical processing following standard protocol available for different reporter gene/s with ease and confidence. Availability and use of green fluorescent protein (GFP) and its other variants as reporter gene/ s to aid in visually determining the expression of the transgene/s into the recipient system has proved essential for study of transgene/s expression. However, use of such reporter systems have proved to be detrimental as they suffer from drawbacks, which warrants discovery and use of alternative reporter genes and elegant precise leak-proof techniques for efficient diagnosis of genetic transformation. Dual-labeling systems, which supercede the efficiency of classical reporter gene/s seem to be more efficient in detecting genetic transformation diagnosis visually. The present study encompasses the cloning and expression of DsRed2 via expression vector (pKYLX80), introgression of gfp into the expression vector and the subsequent introgression of the reporter gene/s i.e., both DsRed2 and gfp into tobacco plant system through microprojectile bombardment to fabricate an efficient dual-labeling framework involving tobacco leaves in non-destructive mode of (non-invasive) reporting of the integration of transgene/s.

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