Abstract
The development of efficient tissue culture protocol for somatic embryo would facilitate the genetic modification breeding program. The callus induction and regeneration were studied by using different parameters i.e., auxins, cytokinins, and desiccation treatment. Scanning electron microscopy and histological analysis were performed to identify the embryogenic callus for regeneration. The callus percentage results showed that MS (Murashige and Skoog) basal medium supplemented with 3 mg/L 2, 4-D and 30g/L maltose were the optimal callus induction medium for MR220 (80%) and MR220-CL2 (95%). The morphology of the embryogenic callus was confirmed by the SEM (Scanning Electron Microscopy) (presence of extracellular matrix surface network) and later by histological analysis. Finally, MS media supplemented with 0.5 mg/L NAA (Naphthalene Acetic Acid), 2 mg/L kin, and 1 mg/L BAP were selected as the optimum regeneration media treatment while callus desiccated for 48 h was proved to produce more plantlets in MR220 (60%) and MR220-CL2 (73.33%) compared to control treatment (without desiccation). The protocol presented here showed the necessity for the inclusion of partial desiccation as an important step in the tissue culture protocol of Malaysian indica rice genotypes in order to enhance their regeneration potential.
Highlights
Successful engineered breeding programs have relied on the establishment of efficient regeneration protocols as well as a selection of efficient DNA delivery method, selection, and maintenance of positive transformants [1]
MS media supplemented with 0.5 mg/L naphthalene acetic acid (NAA) (Naphthalene Acetic Acid), 2 mg/L kin, and 1 mg/L BAP were selected as the optimum regeneration media treatment while callus desiccated for 48 h was proved to produce more plantlets in MR220 (60%) and MR220-CL2 (73.33%) compared to control treatment
The callus induction percentages were increased with the increment of 2, 4-D
Summary
Successful engineered breeding programs have relied on the establishment of efficient regeneration protocols as well as a selection of efficient DNA delivery method, selection, and maintenance of positive transformants [1]. There are a good deal of rice literature [2,3,4] that strategize such protocols to be used in genetic modification studies; successful transformation protocols are severely genotype-dependent [2]. Previous research reported that optimal factors for differentiation and regeneration of indica rice in vitro were pre-requisites before any genetic transformation programs were employed [3]. Visual observation using color-base methods has been used to determine embryogenic rice calli; these may contribute to the misidentification of features and appearance. Determination of embryogenic callus at early callogenesis stages was more effective and reliable in order to minimize the error when selecting the potential of embryogenic
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