Anther culture in cocoa
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Date
1992
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Department of Agricultural Botany, College of Horticulture, Vellanikkara
Abstract
Investigations on anther culture of cocoa (Theobroma cacao L.) were carried out at the College of Horticulture, Kerala Agricultural University, Vellanikkara, during 1990-'92 with the objective of studying the amenability of cocoa anther under in vitro conditions for the production of haploids.
The fertility and viability of pollen grains of cocoa were found to be 35 and 61 per cent respectively.
Anthers at tetrad stage (bud length 1.75 mm) were subjected to a two-stage culture procedure involving incubation and subculture (after 4 weeks) on modified H3 basal medium supplemented with 1 mg 1-1 NAA and 0.1 mg 1-1 2-iP for 50 days (stage 1) and subsequent weekly transfer to ½ Ms basal media supplemented with 0 to 1 mg 1-1 2-iP and 0 to 3 mg 1-1 GA3 (stage II). Proembryoids were obtained in stage 1 medium via callus within 45 days of culture, only when anthers derived from Criollo, Trinitario and H2 were cultured. Serial subculturing of embryoids in stage II media led to formation of shootlets and rootlets. With four sub- cultures in stage II in a span of one month, the embryoids could be germinated into plantlets of size 2.5 cm with two leaflets and one rootlet. This is the first report of plantlet recovery from cocoa anthers via indirect embryogenesis. The plantlets were dried up after an elapse of 3 months. The most favourable combination for root development was Ms medium supplemented with 2 mg 1-1 2-iP, 0.1 mg 1-1 NAA and 126 mg 1-1 PG.
The factors influencing anther callus induction were: stage of anther development, minimum temperature in the field of donor plants, type and strength of basal medium, type and concentration of auxins and cytokinins, carbohydrate source, sucrose level, presence of CW and PG, physical environment and gamma and UV rays. Anther callus multiplication was influenced by type of basal medium; type and concentration of growth regulators; carbohydrate source; sucrose level; presence of amino acids; organic supplements; ethylene releasing and inhibiting chemicals; adenine and its derivatives; unusual regulants.; gibberellins and growth inhibitors; light and gamma rays. The factors influencing indirect embryo- genesis were found to be stage of anther development and light.
Hybrid genotype (H2 and H1) responded more favourably to callus induction, callus multiplication, callus rhizogenesis and were stable with respect to seasonal changes. However, they took more number of days for callus initiation. Criollo took minimum period for callusing.
Chlorine water treated for 3-4 min was the best chemical identified for surface sterilization of flower buds of cocoa. A single auxin or cytokinin alone in the basal medium could not initiate anther callusing. Thermic shock pre-treatments were ineffective for callus induction. Light had no role in callus induction. UV rays had no role in altering anther callus growth. Time of subculturing in cell suspension culture was found to be 15 days. Calloids developed in cell suspensions were not amenable to organogenesis or embryogenesis. Isolated microspore culture was unsuccessful.
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PG
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Citation
170318