Studies On Tissue Culture Of Grimmia Pilifera

Grimmia pilifera belongs to the Grimmiaceae Grimmia, Small xerophytic mosse withstrong characteristics of drought resistance. Taking the gametophyte of Grimmia pilifera asmaterial in this study, the experiment disinfected the explants with the of tissue culturemethod by swirl oscillator. The study explored the sterilize solution, concentration andsterilizing time and getted the aseptic protonematata successfully. The culture conditions ofprotonematata were optimized and the suitable culture medium type for protonematata growth,the concentration of NH4NO3in the culture medium, the species and concentration of sugarsource, illumination, temperature, pH value, the categories and concentration of hormonewere screened. On this basis, the protonematata were induced and the regeneration ofgametophyte were obtained successfully. The main results of this study are as follows:(1) The material can be disinfected effectively through fifteen days of preculture,pretreatment, taking about5mm of the stem apex buds, water rinsed for5h, mercuric chloridesolution of0.025%acuteness oscillated for105s through the aid of swirl oscillator and sterilewater irrigation.(2) Beneck culture medium is suitable for the sterilized Grimmia pilifera explants(gametophyte) returned to green growth.(3) Prat culture medium is suitable for Grimmia pilifera protonematata growth.Beneckculture medium is suitable for Grimmia pilifera protonematatata transverse dispersion growthwith shorter length protonematata. Knop medium is not suitable for Grimmia piliferaprotonematata growth with the black protonematata till death.(4) Prat culture medium with500mg/L concentration of NH4NO3is the most beneficialto the protonematata growth.(5) MS culture medium with sugar and MS culture medium with no sugar have littleinfluence on the protonematata growth;Prat medium with Sucrose or fructose or glucose is all not suitable for Grimmia piliferaprotonematata growth with the black protonematata till death.Beneck medium with Sucrose or fructose or glucose is all not suitable for Grimmiapilifera protonematata growth with the black protonematata till death.(6) When the temperature is (15+1)℃or (19+1)℃, Grimmia piliferaprotonematata grow slowly;(23±1)℃is suitable for Grimmia pilifera protonematata growthand gametophyte regeneration.(27±1)℃is not suitable for Grimmia pilifera protonematatagrowth with the black protonematata and serious browning. (7)3000lx of Light intensity and12h/12h of light cycle are suitable for Grimmiapilifera protonematata growth; Dark conditions make Grimmia pilifera protonematata todeath; all light conditions make Grimmia pilifera protonematata seriously browning andultimately death.(8) PH5.5for Prat culture medium is the most suitable for Grimmia piliferaprotonematata growth; pH6for Beneck culture is suitable for Grimmia pilifera protonematatagrowth.(9) NAA (1mg/L) is not suitable for Grimmia pilifera protonematata growth. KT,6-BA,2,4-D, IBA, IAA have certain stimulative effect on Grimmia pilifera protonematata growth,but the effect among different hormones for Grimmia pilifera protonematata growth areinsignificantly; When the hormone concentration are one of500mg/L,1000mg/L,2000mg/L, the protonematata growth did not change significantly, hormone concentration haveinsignificantly effect for the protonematata growth.(10)Exuberant protonematata have advantageous to new gametophyte growth.(11)MS culture medium with glucose of30g/L could induce gametophytes. But thegametophyte is very weak with poor growth status.(12)Subculturing the browning, yellowing protonematata or protonematata with rhizoidgrowth on the Prat medium of pH5.5, and gametophyte will be grown after20days in goodcondition.(13) Ammonium concentration played an important role on the gametophyteregeneration; Prat culture medium could induce Grimmia pilifera newborn gametophyte;Modified Prat (NH4NO3concentration change from2.5g/L to0.5g/L) can’t induce newborngametophyte.(14)Hormones ratio of certain concetration (2,4-D and6-BA) have poor effect onnewborn gametophyte induction.2,4-D1mg/L+6-BA2mg/L+Prat culture medium caninduce weak newborn gametophyte;2,4-D1mg/L+6-BA1mg/L+Prat medium can induceweak newborn gametophyte, but the Stem and leaf differentiation is not obvious...