STUDY OF PROTOPLAST FUSION BETWEEN Vigna radiata (L.) Wilczek var. radiata AND ITS WILD RELATIVE Vigna radiata (L.) Wickzek var. sublobata (Roxb.) Verdcourt

• Main Author: Angraini (NIM 20698048), Wiwid
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/9637
• Institution: Institut Teknologi Bandung

ABSTRACT: <br /> <br /> <br /> Various PEG solutions were used to induce protoplasts fusion between hypocotyl protoplast Vigna radiata var. radiata and leaf protoplast Vigna radiata var. sublobata, with purposes to find the highest fusion product, and the early stage of development of fusion product and its parents protoplasts were then observed. Protoplasts from hypocotyl of V. radiata var. radiata and leaf V. radiata var. sublobata were isolated enzymatically with enzyme solutions consisted of 2% cellulase YC, 1% macerozyme R-10, 0,4 M mannitol and 2.5 mM CaC12.2H20. The protoplasts of hypocotyl and leaf were fused by using various solutions of polyethylene glycol (PEG) MW 4000 and other compounds namely P1, P2, P3. The highest yield of fusion was 15,27% obtained from P2 solution, which was consisted of 33% PEG, 1,8% sucrose, 1mM KH2PO4 and 10 mM CaCl2.2H20. Observation of nuclear of fusion product which was stained by 4-6 diamino 2-phenilindole (DAPI) under fluorescence microscope indicated that 58,82% uninucleate, 18,23% binucleate, and 22,95% multinucleate. The viability of fusion products high at the beginning of culture until first division. The surface of hypocotyls and leaves protoplasts hail been covered with cell wall material completely after 8 hours of culture, whilst in fusion product after 24 hours. The first cell division was observed after 2 days of culture in hypocotyl and after 3 days of culture in leaf protoplast as well as in fusion products. The capability of fusion product to divide and to form colonies was smaller (0,32%) than hypocotyls protoplast (3,11%) and leaf protoplast (4,28%). Based on the results, it is concluded that the highest protoplast fusion product was induced by P2 solution and the fusion product was able to regenerate cell walls and to form microcolonies.