Breeding System And Pollen Preservation In Vitro Of Paphiopedilum Dianthum

Paphiopedilum dianthum,also called lady slipper,their peculiar shapes bring strong ornamental value.Due to excessive mining by human beings and adverse environmental conditions,the population of p.dianthum has declined sharply and its survival is threatened.Therefore,conservation of p.dianthum requires immediate attention.To provide a theoretical basis for the protection of p.dianthum germplasm resources.The results are as follows:1.Paphiopedilum dianthum blossomed from June to September by observing and recording.Duration of single flower lasts 45 to 55 days and flowering period of single plant was about 75 to 90 days.Each plant generally produced one peduncle bearing with 2～5 flowers.Based on in vitro germination method,p.dianthum pollen viability was measured at different flowering stages.At the blooming period(flowering 5～34d),it reached above 80%and stigma receptivity reached the highest level.The pollinia were persistently located at both sides of the staminode and their position remained unchanged relatively and the hybridization index(OCI)of P.dianthum was 4.The breeding system model was established by artificial controlled pollination experiment.The seed setting rate of self-pollination is 0.Self-pollination cannot be carried out without the help of pollinating media.The seeds setting rates of artificial self-pollination,cross-pollination of the same plant and cross-pollination of different plants were 83.30%,90.00%and 96.70%,respectively.There was no significant difference in these results,which were significantly higher than that of natural pollination.P.dianthum with highly passive self-cross and out-cross abilities,and breeding system requires self-pollination and outcrossing of pollinators.2.Seed aseptic seeding procedure of P.dianthum:P.dianthum matural pod(pollination 300d)with 75%ethanol disinfection 30s+0.1%HgCL2 15min+sterile water 5 times,comes the pollution rate of 0;1/2 MS+100 mL/L Coconut milk+20 g/L sucrose+5 g/L Agar was used for seeds germination comes the best result of evenly full and healthy formed observed at the 50th day.3.Selecting a best method for determination of P.dianthum pollen viability,research into the effects of different storage methods on pollen life,and explore the cryopreservation procedure.(1)Pollen germination was best when cultured in liquid medium containing 5%sucrose+0.05%boric acid+ 0.001%calcium chloride for 36h in vitro,which was the most accurate method for the determination of pollen viability.(2)The life span of the pollen was less than one week at 27℃,and the pollen viability decreased with the increase of storage time at 4℃ and-20℃.When the pollen was stored for 5 days,the order of pollen viability is-196℃,4℃,-20℃,27℃.The lifespan of the pollens stored at ultra-low temperature(-196℃)was the longest,and the germination rate was still up to 69.86%when stored for 16 days.(3)Procedure of pollens cryopreservation:fresh pollens(moisture content 31.8%)were immersed into LN without drying dehydration,and thawing by room temperature 30min,running water 15min or lmin at 37℃ water after cryopreservation.