Study On Somaclonal Variation Of Zoysia Matrella

Zoysia matrella(L.) Merr.,namely manilagrass,is an excellent warm-season turfgrass species,which is widely used in golf courses,ground track fields and parks.As Zoysia matrella produces asexually by stolons in nature,conventional hybridization breeding procedure cannot be applied for its genetic improvenment.Thus modern biological technology such as somaclonal variation is an effective way to increase its resistance. In this experiment,embryogenic calluses induced from the stolons of manilagrass were taken as materials.Effects of different types of culture(suspension culture and solid culthure),different basal media(MS,N6,N6AA) and concentrations of 2,4-D (0.5,1.0,1.5,2.0 mg·L^-1),different concentrations of sucrose(1.0%,2.0%,3.0%,4.0 %,5.0%),different kinds of cytokinins(blank control,BA,KT,ZT,TDZ),different culturing temperatures(4℃and 25℃),different subculture periods(1 year and 5 years),different doses of ⁶⁰Coγ-irradiation(0,5,10,20,40 Gy)on calluses growth, regeneration,physiological and genetic stability were compared.Calluses were screened on media with different concentrations of NaCl(0%,1.0%,1.5%,2.0%) and glyphosate(0,0.01,0.02,0.05,0.1,0.5,1.0,1.5,2.0 mmol·L^-1).The primary results were summarized as follows:1.Compared with solid culthure,growth rate of calluses was increased significantly through suspension culture,but regeneration capacity of callus was decreased. Therefore,it was more suitable for short-term rapid proliferation of calluses by suspension culture.The optimum basal medium for calluses subculture of manilagrass was MS medium.Calluses could maintain vigorous growth,well embryogenesis and high regeneration capacity on MS medium with 2.0 mg·L^-1 2,4-D and 40 g·L^-1 sucrose. BA was suitable for calluses regeneration.Calluses on the regeneration medium with 0.1 mg·L^-1 BA could differentiate and form more plantlets with high rooting ability, which ensured the survival of the regenerated plants and improvement of the activity of catalase(CAT) after transplantation.2.Regeneration rate of the calluses maintained at 91.7%-97.9%during 11 months' low temperature culture at 4℃.But the speed of calluses regeneration was decreased and the duration of regeneration was prolonged after the low temperature treatments. Regeneration capacity of the calluses that had been cultured for 13 months at low temperature declined significantly.Regeneration rate of the calluses subcultured for 5 years showed no significant variations compared with that subcultured for 1 year.3.⁶⁰Coγ-irradiation at lower dose(5 Gy) had no effects on growth and regeneration of the calluses.Middle doses of ⁶⁰Coγ-irradiation(10 and 20 Gy) had a significant promoting effect on regeneration of the calluses.Higher dose of ⁶⁰Coγ-irradiation(40 Gy) resulted in the decreases of embryogenic maintenance ability and regeneration rate of the calluses.Proline contents in the regenerated plants acquired from the calluses that were treated with ⁶⁰Coγ-irradiation at the doses of 5,10,20 and 40 Gy were significantly higher than that in control(0 Gy),and the highest content(20 Gy) was 2.4 times of that in control.4.⁶⁰Coγ-irradiation at the dose of 10 Gy resulted in the increases of catalase(CAT) activity and proline content in the regenerated plants by 16.0%and 1.2 times, respectively.And there was a remarkable increase in the regeneration capacity of the calluses on medium with NaCl.9.6%of the calluses treated by 10 Gy ⁶⁰Coγ-irradiation regenerated and formed plantlets on the regeneration medium with 1.0%NaCl.The regenerated plants grew vigorously after being explanted to rooting medium with 1.0%NaCl.5.After 3 months' screening,embryogenic rate of the calluses of manilagrass still reached 45.0%even on the subculture medium with 2.0 mmol·L^-1 of glyphosate. These resistant calluses maintained a high regeneration rate of 100%.However, calluses were extremely sensitive to glyphosate when they were regenerated.There were no regenerated plants acquired on the regeneration medium with glyphosate.