Morphological and histological analysis of anther-derived embryos of coconut (Cocos nucifera L.)

Abstract

Doubled-haploids are a great source of material for heterozygous coconuts to shorten the time taken for crop improvement through hybridization programs. However, low frequency of embryo conversion and formation of weak plantlets are the major limitations in the protocol. In the present study anther-derived embryos were analysed at cellular level through the histological observations to understand the occurrence of shoot differentiation in those embryos. Attempts were also made to optimise the 6-Benzylaminopurine (BAP) concentration in the regeneration medium. Among the tested BAP levels (5, 15, 25, 35, 45, 55 μM), the regeneration medium containing 25 and 35 μM gave rise to 51.5% sprouted embryos against the control that gave only 22.7%. Among the analysed anther-derived embryos sequential events of differentiation including the formation of provascular strands followed by vascular bundle, differentiation in to the growing point, giving rise to the secondary embryos and polarization in to the shoot and root pole were identified. The blunt embryos devoid any morphological sign of sprouting showed different cellular arrangement viz. haustorial structure without any meristematic point (47%), bipolar with shoot and root meristems (8%) and unipolar with either of the pole (45%). Thick haustorial cover lead to the physical dormancy inhibiting further development of the polar structures. The germinating embryos containing a single shoot gave rise to the healthy plants where as double, multiple or fused shoots formed the week plantlets. The findings can be used for further optimisation of the protocol to achieve a greater plant regeneration frequency. 1. Culture media supplemented with increased levels of 6-Benzylaminopurine (BAP) enhanced the regeneration frequency of anther-derived embryos. 2. Thick haustorial tissue created an obstacle effect on further development of the differentiated shoot apical meristem. 3. Formation of clustered plants through secondary embryogenesis or shoot multiplication at the early stage of plant regeneration caused to give rise to the week plants.