Polyploidy Induction And Identification Of Wild Mulberry Germplasm Resources

Mulberry is a perennial deciduous plant and one of important economic trees in China.Polyploid mulberry has high yield with high quality and strong resistances.Most of the studies on the induction of polyploidy have been mainly performed using the mulberry cross-breeding seedlings or cultivated varieties,such as the tetraploid mulberry was induced to octoploid material,and crossed with another tetraploid mulberry to produce a hexaploid mulberry in order to improve the production of mulberry leaves.Wild mulberry resources as a material for mulberry polyploid induction has not been reported yet.The varieties among the different ploidy materials in morphological and biochemical characteristics have functional significance for breeding and provide valuable information to elucidate the mechanisms of plant polyploid evolution.The adventitious bud of wild mulberry resource Yun 7(2n=7x=49)and Morus notabilis 2n=2x=28)was used as experimental materials in this study.The induction of adventitious bud and rooting were performed.The effects of concentration and treated time of colchicine on the induction of polyploidy of Yun 7 and M.notabilis were also investigated.To obtain homozygous fourteen-ploidy(2n=14x=98)and tetraploid(2n=4x=28)mulberry,mixoploids were subsequently screened and the axillary buds were finally separated.Fourteen-ploid and tetraploid mulberrywere obtained.Themorphological and photosynthetic characteristics of Yun7 and its mutagenic plantswere measured.The artificial induction of chromosome doubling in wild mulberry was successful.The results are as follows:1.Polyploidy induction of germplasm resource No.7 of Morus wittiorum Hand-MazzThe adventitious bud induction system of wild mulberry resources Yun7 was established.The adventitious buds were induced by winter bud.Induction of adventitious bud medium: MS + 30g/L Sucrose + 1.5 mg/L 6-BA + 100 mg/L Inositol + 8 g/L agar.For rooting medium: MS+ 30g/L Sucrose + 0.5 mg/L NAA + 0.4 g/L activated charcoal + 8 g/L agar.The adventitious buds were soaked in 1 g/L,2 g/L,and 3 g/L colchicine for 2 days,2.5 days,and 3 days,respectively,to induce of polyploidy of Yun 7.The results indicated that 2 g/L colchicine combined with dimethyl sulfoxide used as the penetrant and treated for 2.5 d,the half-death rate of cultured seedling were up to 53.76%,which had optimal effect for the polyploidy induction.To obtain homozygous fourteen-ploidy mulberry,resultant mixoploid was further screened and the axillary buds were continuously separated.Finally,the fourteen-ploid mulberry(2n=14x=98)was obtained.2.Trait measurement of Yun 7 and mutagenic plants(2n=14=98)Leaf morphology,photosynthetic efficiency,and other properties of Yun 7 and mutagenic plants were determined.The results showed that in mutagenic plants,the leaf became larger and their edges were deeper.There were significant differences between the mutant plants and Yun 7 in leaves.The mutagenic plants grew faster and higher with larger stomatal cell.The photosynthetic rate of mutagenic plant was increased.As a summary,the mutagenic plants were grown well.3.Polyploidy induction of M.notabilisThe adventitious bud induction system of wild mulberry resources M.notabilis was established.Embryo rescue technique was used to make the seeds germinate.The adventitious buds were induced by hypocotyl.Induction of adventitious bud medium:MS +30g/L Sucrose + 1.0 mg/L 6-BA + 8 g/L agar.The adventitious buds were soaked in 0.1 g/L、0.5 g/L and 1 g/L colchicine for 12 h,18h,and 24 h,respectively.The results indicated that the death rate of 1 g/L colchicine treated for 12 h and 18 h were 43.58% and 57.58%,respectively.In these conditions,the tetraploid induced by M.notabilis was obtained.The polyploid induction of seed germination was carried out immersion method.The germinated seeds died.Using titration method,axillary buds were induced and grew normally but,no mutagenic plants were detected.In this study,we used the mulberry tissue culture technology to solve the problem that the Yun 7 and M.Notabilis were not easy to reproduce asexual.The adventitious bud induction system of Yun 7 and M.notabilis was established to promote the genetic transformation,preservation andexploitation of wild mulberry germplasm resources.The artificial induction of chromosome doubling in wild mulberry was successful.Differences of ploidy materials in morphological and biochemical properties have functional significance in polyploid breeding.The present study provided valuable materials(fourteen-ploidy and tetraploid mulberry)to better understanding the polyploidy of plant and facilitate the study of the mulberry origin and evolution.