Establishment Of A Tissue Culture System For "Huaye 5" Oryza Alta Swallen Line And Induction Of Autoallopolyploid

Oryza alta Swallen is native to tropical regions of South America.It contains many excellent genes that are not available in cultivated rice and is a valuable resource for rice breeding.However,the Oryza alta Swallen chromosome group was CCDD,which had a long-distance relationship with the cultivated rice of AA group.Hybrids are prone to infertility,which made it difficult to carry out favorable gene transfer and hindered the research and utilization of Oryza alta Swallen.Studies have shown that homologous polyploids can overcome infertility.Therefore,this study intends to use the strain “Huaye5”,which was selected from the offspring of Oryza alta Swallen,as a material to establish a Oryza alta Swallen doubling system and induce the production of homogenic and heterogenic Oryza alta Swallen.The main research contents include: First,the establishment of the tissue culture system of the "Huaye 5" Oryza alta Swallen line;Second,the use of the obtained calli,doubled by colchicine to obtain autoallopolyploid Oryza alta Swallen.Third,observe and study the main traits of "Huaye 5" Oryza alta Swallen line with different origins.Here are main results:(1)Establishment of a "Huaye 5" Oryza alta Swallen line tissue culture system.By comparing different basic medium,hormone ratio and differentiation medium,experimental studies were carried out to establish a suitable culture system for this material: The mature seed of Oryza alta Swallen was used as the material,and the appropriate program for callus induction was MB +2,4-D(2.5mg/L)+ 6-BA(1.0mg/L),the average induction rate was approximately 15.63%;Subculture medium was: NB+2,4-D(2.0mg/L)+0.2g/L activated charcoal;the suitable protocol for callus differentiation is MS+KT(2.0mg/L)+NAA(0.2mg/L).The average callus rate under this protocol is about 10.15 %.(2)Induction of autoallopolyploid of "Huaye 5" Oryza alta Swallen line.Colchicines were soaked in callus and co-cultured to treat young shoots and callus in two ways.Thefollowing results were obtained: First,the callus was soaked with an aqueous solution of colchicine at concentrations of 200 mg/L,400 mg/L,600 mg/L,and 1000 mg/L,respectively.After treatment for 24 hours,the regenerated seedlings grew at an average of6.75% for the four treatment groups,of which 600 mg/L had a higher seedling emergence rate of 12.96%.No re-differentiated seedlings were produced in the colchicine treatment at48 hours.The second is that the co-culture method for the treatment of callus and young shoots has a colchicine concentration of 200 mg/L and is treated for 120 h.Among them,the callus regeneration rate after treatment was approximately 5.00%;the final seedling rate of young cluster buds was approximately 30%,and a autoallopolyploid Oryza alta Swallen was detected.The variation rate among the obtained seedlings was approximately at66.67 %,the induction rate of the final autoallopolyploid was approximately 5.00%.(3)Morphological characteristics of autoallopolyploid of "Huaye 5" Oryza alta Swallen line.The number of chromosomes in the root tip indicated that there were 48 chromosomes in the control plants and the number of chromosomes in the autoallopolyploid plants was 96.Compared with the original species,the plant height of "Huaye 5",with chromosome doubling,was significantly reduced by 19.31%.leaf length and leaf width were both significantly increased,with increases of approximately 74.71%and 29.71%,respectively.The number of childbirths decreased significantly by about79.00%.Observing the stomata on the surface of the leaf,the stomatal density on the leaf surface of the autoallopolyploid plants was reduced and the volume was increased.Through the above studies,a tissue culture system suitable for the "Huaye 5" Oryza alta Swallen line was built,and a certain number of Oryza alta Swallen autoallopolyploid was obtained through colchicine chemical mutagenesis methods,confirming and preliminary understanding.At the same time,some agronomic traits of the material were investigated to provide basic material protection for follow-up research and breeding.