| Pea(Pisum sativum L.) is a leguminous herbs of a year or more, is an important food, fodder, vegetables, fertilizer and medicine crop. In recent years, peas play an important role in the development of animal husbandry and sustainable agriculture, Planting area and market demand are growing. However, due to the impact of pests and drought, cold and other adverse conditions, the yield is very unstable. Establishment of regeneration system by plant tissue culture and studies on genetic stability will provide the foundation for in vitro regeneration and genetic transformation.The semi-leafless peas cultivar ?Longwan NO.1? and vining peas cultivar ?S3008? were chosen as material. Used to study the effect of different genotypes, hormone types and the ratio of medium on callus induction and callus differentiation. The research results are fllowed:1.By comparing the different explants and culture conditions,the best explant is pea stem cultured in dark. the optimal medium for callus induction was MS+2 mg·L-1TDZ+0.2mg·L-12,4-D, The frequency of callus initiation from stems of ?Longwan NO.1? and ?S3008? was 88.7% and 86.7%. The explants were cultured on MS only with TDZ to induce, callus can not formate or formate slowly, the color become lighter and yellowed eventually. Lower concentrations of 2,4-D can significantly improve the callus induction rate, It is not conducive to adventitious buds differentiation with high concentrations of 2,4-D to induce, callus growth vigorous and callus browning eventually, lots of white granular protrusions on surface.2.The appropriate medium for adventitious bud induction was MS+2 mg·L-1TDZ +1mg·L-16-BA. The frequency of differentiation of ?Longwan NO.1? and ?S3008? was 76.7% and 74.7%. the adventitious bud number per explants was 4.2 and 4.4. TDZ compare with 6-BA can promote adventitious buds differentiation. Under the same hormone concentrations, both the adventitious bud regeneration rate and the average number of adventitious buds, TDZ was better than 6-BA.3.Through study the inhibit effect of different antioxidants and sorbents on callus browning, we found out the best medium for inhibition of callus browning is MS+1.5 mg·L-16-BA+0.2mg·L-12,4-D+0.05g·L-1 riboflavin, The two varieties callus Browning rate reduced to 18.0% and 20.7%, better and quicker growth. When added 0.5g·L-1VC into the medium, the callus browning rate decreased to 28.7 and 34.0% Significantly lower than 0.1 and 0.3g·L-1VC. Culturing in the medium with 0.5g·L-1AC the browning rate of callus is 51.3% and 50.7%. When the concentration of AC was 1.5g·L-1 browning rate reduced to 34.0% and 35.3%, but the callus growing weaker.4.The best medium for adventitious buds subculture was MB5+1mg·L-16-BA +1mg·L-1IAA,two different kinds of adventitious buds were observed depending on the presence or absence of GA3 in the medium.. The adventitious buds produced on media containing GA3 was lighter and more vimineous, compared to adventitious buds cultured on medium without GA3.5.Through study on the impact of stem explants directly induce adventitious buds, The appropriate medium for stem explants directly induce adventitious buds was MS+4mg·L-1TDZ+0.3mg·L-1IAA. ?Longwan NO.1? and ?S3008? adventitious buds differentiation rate were 78.7% and 80.0%. The average number of adventitious buds more than 3.6.The adventitious buds on rooting induction medium(1/2MS+20g·L-1 sucrose+1.5mg·L-1ABT), the rooting rates was 74%, The average number of adventitious rooting was 4.2.7.By chromosome analysis, Two varieties of pea chromosome number of the regenerated plants were same to the donor plants(2n=2x=14), which suggested that the regenerated plants were genetic stable. |
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