Induction of heteroploidy in Dendrobium sp

Abstract

The present study entitled “Induction of heteroploidy in Dendrobium sp.” was carried out at the Department of Plant Breeding and Genetics, College of Agriculture, Vellayani during 2019-21. The study was undertaken to induce heteroploidy in Dendrobium crumenatum using antimitotic agent colchicine via in vitro and in vivo technique. In in vitro induction of heteroploidy in Dendrobium crumenatum, the explant used in the micropropagation was stem nodal segments containing one axillary bud and the basal medium chosen was VW medium. The best combination of surface sterilization agents identified were, 20 per cent sodium hypochlorite for 20 minutes followed by 0.1 per cent mercuric chloride for 5 minutes and 70 % alcohol for 2 minutes. The medium VW+1 mgL-1 BA + 0.5 mgL-1 2,4-D was chosen for callus induction, callus proliferation and PLB formation. The explants were maintained in dark for callus induction and proliferation and for subsequent stages a photoperiod of 16 hours light and eight hours dark was provided. For the induction of heteroploidy, PLBs were subjected to eight different colchicine treatments in basal VW suspension medium. Colchicine treatments were based on the concentration of colchicine (0.05%, 0.10%, 0.15%, 0.20%) and duration of exposure of PLBs to colchicine (24 and 48 hrs). For shoot induction, VW basal medium supplemented with the hormones BA (1mgL-1) and NAA (0.1 mgL-1) was used. The maximum survival percentage of cultures at shoot multiplication stage after different colchicine treatments was observed to be 83.33 per cent, and the minimum survival percentage noted was 25 per cent. In in vivo induction of heteroploidy in Dendrobium crumenatum, D. crumenatum plantlets were subjected to eight different colchicine treatments, based on colchicine concentration (0.05%, 0.1%) and the duration of exposure of plantlets to these doses of colchicine (24 hrs, 48 hrs, 72 hrs, 96 hrs). The highest percentage of plants showing response after colchicine treatment was found to be 83.33 per cent (observation recorded after 2 weeks of colchicine treatment) and the lowest percentage recorded was 16.67 per cent. The observations shoot length, diameter of pseudobulb, number of leaves and width of leaves showed mean values greater than their corresponding control means, whereas the observations, diameter of shoot and length of leaf noted lower mean values than corresponding control. The mean value for the character height of pseudobulb were found to be same for treated and control plantlets. The character, number of leaves recorded high PCV and GCV. Moderate PCV and GCV were recorded for the characters viz., diameter of pseudobulb, length of shoot, length of leaf and width of leaf. The characters, diameter of shoot and length of pseudobulb obtained low PCV and GCV. Significant positive genotypic correlation was observed between length of shoot and height of pseudobulb, diameter of shoot and width of leaf, length of pseudobulb and number of leaves, number of leaves and length of leaf and width of leaf and number of leaves. Flow cytometry analysis of the colchicine treated in vivo and in vitro D. crumenatum samples were performed at Rajiv Gandhi Institute for Biotechnology, Poojapura. The histogram peaks obtained after the analysis of control (diploid) and treated samples were compared for interpretation of the results. In the in vitro treated samples, the maximum tetraploids (66.67 %) were obtained from treatments 0.15% colchicine, 24 hrs and 0.10 % colchicine, 48 hrs. On analyzing the histogram peaks of in vivo treated samples, the highest tetraploid induction (50%) was noticed in treatment 0.05% colchicine, 96 hrs. It was concluded from the present investigation that, for Dendrobium crumenatum which was subjected to both in vitro and in vivo colchicine treatments, in vitro colchicine treatment was found effective than in vivo colchicine treatment, when the concentration and duration of exposure to colchicine used for the study were same.