5.1. Studies on in vivo shoot multiplication in hypocotyls of Rhizophoraceae mangroves: a new approach for scaling up of vegetative propagation
- To evaluate Species-specific capability of multiple shoot regeneration (in vivo) in Bruguiera parviflora, B. gymnorrhiza, Rhizophora apiculata and Kandelia candel
- Biochemical and anatomical analysis during shoot induction process in Bruguiera parviflora, B. gymnorrhiza, Rhizophora apiculata and Kandelia candel
- Adventitious Rooting in multiple shoots of B. parviflora and B. gymnorrhiza
- Hardening of rooted saplings of B. parviflora and B. gymnorrhiza
- Interpreted organogenic development (histological basis) of multiple shoot regeneration in all the target species i.e. Bruguiera gymnorrhyza, B. parviflora, Kandelia candel and Rhizophora apiculata.
- Documented changes in Protein Profile (gel-electrophoresis) during multiple shoot regeneration in hypocotyls of B. parviflora, Kandelia candel and Rhizophora apiculata.
- Standardized method of regeneration of adventitious rooting in multiple shoots of B. parviflora and B. gymnorrhiza
- Saplings raised from vegetative propagation were supplied to mangrove forest division, Bhitarkanika for establishment trial in wild recently.
Performance of Multiple shoot regeneration
One month old established propagules of Bruguiera gymnorrhiza, Bruguiera parviflora, Kandelia candel and Rhizophora apiculata belonging to the family Rhizophoraceae were decapitated following standardized method to induce multiple shoots and species-specific ability was recorded.
Amongst four studied species, the best shooting ability was recorded in the hypocotyls of Kandelia candel. Around 85-95% of the hypocotyls gave rise to multiple shoots with an average of 5.5 nos per hypocotyl regenerated within 25-35 days followed by B. gymnorrhiza , B.parviflora and R.apiculata.
Organogenic development of multiple shoot initiation in hypocotyls
By day 20 of decapitation, shoot primordia were found initiated from meristem cells in the region of the severed vascular bundles near the exposed surface of the hypocotyls. Decapitation-induced meristematic cells gave rise to vascular-connected multiple shoots by day 45.
Formation of shoot/bud primordia with tunica layer observed at day 20 (long arrow). Well formed shoot initials with vascular connection recorded at day 30 ( arrowhead).
Shoot initials with vascular connection ( arrowhead) at 30th day of decapitaion
Shoot/bud primordia formation with tunica layer noticed at day 20. Fully developed shoot initials possessing vascular connection recorded at day 30.
Development of shoot/bud primordia with dense cytoplasmic cells observed at day 20. Emerging shoot initials having vascular connection noticed at day 30 .
Studies on changes in Protein Profile during multiple shoot regeneration in Kandelia candel, Rhizophora apiculata, Bruguiera parviflora through SDS-PAGE
Protein synthesis is an essential bio-molecular process during cell division and growth. Salinity impairs normal growth and overall development process in plants. Proline directly or indirectly play an important role in protein accumulation and in cell adaptation to salinity stress. The present study was undertaken to examine specific differences in protein patterns due to salinity and/or proline effect on multiple shoot organogenesis through SDS-PAGE following standard method.
In Kandelia candel, organogenesis of multiple shoot witnessed synthesis of 5 polypeptides having molecular weights ranged from 14-29 KDa at different stages of induction affected by salinity stress and proline application.
In Rhizophora apiculata, synthesis of 5 polypeptides having molecular weights ranged from 14-43 KDa were recorded at different stages of induction affected by salinity stress and proline application.
In Bruguiera parviflora, however, induction of multiple shoot triggered synthesis of 6 polypeptides having molecular weights ranged from 14-40 KDa at different stages of induction affected by salinity stress and proline application.
Adventitious rooting in multiple shoots of Bruguiera gymnorrhiza, Bruguiera parviflora and Rhizophora apiculata
An experiment was conducted to optimize rooting in air layering of three tree mangroves species namely Rhizophora apiculata, Bruguiera gymnorrhiza, Bruguiera parviflora using IBA (5000 mg/l), NAA (1000 mg/l) or in combinations. The overall rooting response in terms of percent rooting, root number, rooting time, root length were found better in the treatments with combinations of IBA (5000 mg/l) or NAA (1000 mg/l) as compared to IBA (5000 mg/l) or NAA (1000 mg/l) alone in B. gymnorrhiza and B. parviflora ; R. apiculata, however, did not respond to rooting. Better rooting performance (80%) and mean root number (5.0±1.0), mean root length (2.0±0.10) were recorded in B. gymnorrhiza treated with IBA (5000 mg/l) +NAA (1000 mg/l) (T3). More than 80% of the rooted air layers survived in the nursery in polybags.
Fig. B. gymnorrhiza : A-samplings with multiple shoots; B-rooting (arrow) through layering; C-hardened saplings in polybags.