Study On Optimization Of Pear Subculture System And Its Regeneration

China is one of the birthplaces of the genus Pears.The white pear varieties,sand pear varieties,and Qiuzi pear varieties widely cultivated in production are native to China.The types and cultivation areas of pears rank first in the world.Pears are perennial woody plants based on vegetative reproduction,which have high genotype heterozygosity and long childhood.The research process of pears is slow,which is difficult to improve through traditional breeding methods.The research on genetic transformation technology of in vitro tissue regeneration on pears promotes the development of pear germplasm innovation and the breeding of varieties,which is an effective method to realize genetic engineering breeding of pears.Establishing an efficient subculture system and regeneration system is the basis of genetic transformation research,so it is very important to obtain high-quality and efficient propagation system and regeneration system.Materials,aiming at the problem of malformation of wild Qiuzi pear(Pyrus ussuriensis M.),optimize the subculture system of pear to provide a wide range of experimental materials for subsequent research.And study the factors that affect the effect of transgenic,and establish a stable system of pear genetic transformation system.Cuiguan pears are used as experimental materials to establish a multiplication system and cultivate an efficient Cuiguan regeneration culture system,which lays the foundation for transgenic research.The main findings are as follows:1.This article is to optimize the subculture system of wild Qiuzi pear(Pyrus ussuriensis M.).In view of the abnormalities in the subculture process,the effects of 6-BA and IBA hormone ratio on tissue culture seedlings are studied.The results show that 6-BA mainly promotes the growth and differentiation of plants.With the increase of 6-BA concentration,the bud proliferation rate increases,and the leaves increase,but the concentration is too high,but it inhibits growth and the plant browning is severe.However,the appropriate IBA content promotes the strong growth of the plant,and the leaves are bright green and the plant is healthy.Therefore,the appropriate hormone ratio plays a key role in the growth of the plant.The medium formula of the improved Qiuzi(Pyrus ussuriensis M.):MS+2.5 mg·L-1 6-BA+0.3 mg·L-1 IBA+30 g·L-1 sucrose+ 7.5 g·L-1 agar,the average bud proliferation rate is 2 and the browning rate is 0.In the process of affecting the effect of genetic transformation,the leaf age index,leaf infection time,bacterial concentration,dark culture time and other factors were explored.The study found that:choose leaves with thick texture,healthy growth,and leaf age about 20d Infection,leaf infection for 20min,bacterial concentration 0.8,optimal dark culture time at 18 days:pre-culture 3d,co-cultivation 5d,screening dark culture 10d,regeneration rate can reach 90%,and differentiated adventitious bud Strong,browning is 0.2.Using Cuiguan as experimental material,to study the influence of medium type,hormone ratio and antibiotic type on the subculture system of Cuiguan pear,the best subculture medium formula suitable for the growth of Cuiguan pear is:MS+1.0 mg·L-16-BA+0.2 mg·L-1 IBA+0.1 mg·L-1 TMT+30 g·L-1 sucrose+7.5 g·L-1 agar,the bud proliferation rate was 1.82,and the browning rate was 0.Through the ratio of the medium type and different hormones.Based on the exploration of leaf placement method and dark culture time,the best medium formula for regeneration culture is:NN69+2.0mg·L-1TDZ+0.5 mg.L-1IBA+30 mg·L-1 sucrose+7.5 mg·L-1 agar,dark culture time 21d,leaf inoculated on the medium with the distal axis of the leaf,the regeneration rate can reach 88.9%.The average regeneration rate per leaf was 2.58,the callus regeneration rate reached 100%,and the browning rate was 0.