PRIMER DESIGNING AND STANDARDIZATION OF PRIMER ANNEALING TEMPERATURE FOR BETA-3-ADRENERGIC RECEPTOR IN MUS MUSCULUSAbstract
Adrenergic receptors (ADRs) are implemented in important pharmacological responses. The β3adrenergic receptors have been proposed as a suitable drug target for treating diabetes and obesity. However, very little information is available regarding the expression profile of β3 receptors. We have designed primers for studying the β3adrenergic receptors in C57BL/6J mice using various bioinformatics tools in the present study. The primers were designed using three different software, primer 3 Plus, NCBI Primer BLAST, and Primer express, and validated using UCSC in-silico PCR and NCBI -Primer-BLAST. The best primer pair was selected based on primer melting temperature, GC% and primer length. The selected primer pair was custom synthesized. The primer annealing temperature was standardised using gradient PCR. Among the three software used for primer designing, primer express was found to be the best, and the primers designed from primer express were found to have the ideal melting temperature between 58-60°C, 50% of GC content, and 20bp length. From gradient PCR, the primer annealing temperature was found to be 58.3°C. The bioinformatics tool is useful for the successful designing of primers. Primer express software is best among other software used to design qRT PCR-specific primer.
Vijayalakshmi Gangadhara and Asha Abraham *
Department of Post Graduate Studies & Research in Biotechnology, St Aloysius College (Autonomous) - Affiliated to Mangalore University, Mangalore, Karnataka, India.
17 August 2022
13 October 2022
31 October 2022
01 April 2023