PLANT CELL BIOTECHNOLOGY AND MOLECULAR BIOLOGY
 

PLANT CELL BIOTECHNOLOGY AND MOLECULAR BIOLOGY, ISSN No. : 0972-2025, Vol.: 18, Issue.: 7-8

Original Research Article

STANDARDIZATION OF PROTOCOL AND DETECTION OF BBTV FOR TISSUE CULTURED PLANTLETS OF BANANA Cv. Safed velchi THROUGH SANDWICH ELISA TECHNIQUE

 

S. S. KADAM*, N. B. GOKHALE, S. V. SAWARDEKAR, M. S. JOSHI AND V. G. KELKAR 

Plant Biotechnology Centre, College of Agriculture, Dr. Balasaheb Sawant Konkan Krishi Vidyapeeth, Dapoli 415712, Maharashtra, India [SSK, NBG, SVS, MSJ, VGK]

Abstracts

The present study was carried out with an objective to standardize protocol for viral detection and to test tissue cultured banana plantlets for Banana Bunchy Top Virus (BBTV) through ELISA technique. For detection of BBTV infection, the plant extract was isolated from tissue cultured tender leaf samples of banana plantlets. A total of 200 plantlets were selected for extraction of plant sample from 3 mother plants (mother plant no. 15, 16 and 21). The standardization of buffer constituents for ELISA was used. Modifications in extraction procedure and incubation periods in ELISA procedure showed minimum time and clear colour formation for visual detection, an economic protocol for ELISA has been established for both time and chemicals. It was observed that, only positive control and BBTV infected sample of banana plants showed positive results. While, all the remaining sample test wells showed the negative results. It is also observed that, quality of antigen-antibody complex get increased, minimize contamination by increasing volume of wash buffer up to 400 µl and incubation time for good colour formation. ELISA Assay is an effective technique for the detection of BBTV infection of tissue cultured plants. None of the micropropagated plants gave positive reaction with the antiserum of BBTV. These modifications led to minimum time and clear colour formation for visual detection, showing an economic protocol for ELISA for virus detection in tissue cultured banana plantlets. Therefore, the immunological studies showed the absence of BBTV in the micropropagated plantlets. This test has been made mandatory to all the tissue culture laboratories so that viral/non-viral plantlets will be detected.

Keywords :

Tissue culture, micropropagation, Banana Bunchy Top Virus, ELISA, antibody.