In Vitro Regeneration And Chromosome Doubling Of Spathoglottis Plicata

Spathoglottis plicata is a kind of tropical terrestrial Orchidaceae orchid which has purple flower.It has long florescence and a lot of flowers.lt can be widely used for potted plant, cutting flower and take the place of summer annual flower.And it has a high value of gardening and application value. In this experiment we use spathoglottis plicata as material.The in vitro protocorm-like-bodies(PLBs) rapid propagation regeneration system, the high frequency PLBs induction system and the ployploid induction system of spathoglottis plicata by using colchicines In vitro were studied.Here in,brief results are as follows:1 .A reproducible protocol for protocorm-like-bodies(PLBs) of Spathoglottis plicata was established. Using its axillary bud as explants, the effect of different hormone concentration ratio and additives on PLBs induction, differentiation and rooting were studied. Rapid propagation plantlet obtained after 35d of culture on the half Murashige and Skoog (MS) basic salts supplemented with 2.0 mg/L 6-BA, 0.2 mg/L NAA, 0.2 mg/L AC(activated carbon), 10% CW (coconut water). The survival rate of axillary buds rase up to 80% after cultivation for 35 days. The PLBs clusters induction medium of apical meristem in vitro was consisted of 10 mg/L Picloram in half MS basic salts, and the highest induction rate of PLBs clusters was 75%. The PLBs differentiation percentage of 54% were observed in differentiation media, which composed of half MS basic salts, 2.0 mg/L 6-BA, 0.2 mg/L IBA. The frequency of plantlet regeneration in the rooting medium with the half MS basic salts, 1.0 mg/L IBA, 0.5 mg/L 6-BA, 0.2 g/L AC was 85% after cultured for 35 days. Rooted plantlets were acclimatized with a survival rate of 95% and grew vigorously.2.A high frequency PLBs induction system of Spathoglottis plicata by using tTCL(transverse thin cell layer) was established. The effect of basic culture media,hormone concentration ratio and the aspect of explant on PLBs induction were studied.TCL showed the highest survuval rate by 78.6% on 1/2MS, yellow granular PLBs could be induced and grew well on this media.The PLBs induction rate of shoot point location was 40.6% ,which was higher than young leaves and basis roots.Matured PLBs was obtained after 35d of induction training.tTCL from the location of shoot could had the highest PLBs induction rate as 45.6% on 1/2MS+Picloram 5mg/L+6-BA 0.5mg/L.Mutured PLBs differentiated into shoots on 1/MS without hormone after 40d.3. A ployploid induction system of Spathoglottis plicata by using colchicines in vitro was established. Using its its seed and TCLs as explants, the effect of colchicine concentration ratio and processing time on ployploidy induction were studied. Seeds mixture culture method got a highest mutation rate of 17% in 0.04% colchicine for 20d.Dropping liquid on tTCL got a highest mutation rate of 27.8% by 0.3% colchicine for 30d.191 mutant plant were obtained by morphological observation and stomatic observation.There were 10 pure tetraploid (2n=4x=80) and 38 chimera by chromosome counting identification. The suitable isolation and purification method of S.plicata was tTCL of mutant plant.Obtain 6 pure tetraploid in 13 regenerated plant.