Study On Technique Of Anther Culture In F₁ Between Upland Rice And Lowland Rice And The Establishment Of Doubled-haploid Lines

Doubled-haploid progenies are very useful especially to breed new varieties and construct high density genetic linkage maps, which play important roles in gene localization and gene cloning. An efficient anther culture technique is necessary to produce usable D-H progenies. Although the study of the anther culture of rice was started in early 1970's, many problems remain. For examples, browning and the selection of culture media. In this article, anthers culture of seven genotypes were studied to investigate some important factors affecting the anther culture of in F] between upland rice and lowland rice. The results showed that the frequency of callus induction and differentiation were affected by the genotypes of materials, date of sample colleting, developmental stage of anthers, days of lower temperature pretreatment, formula of culture medium and so on.Better medium formula was the combination of macroelement in N6+microelement and organicelement in SK3+2,4-D (0.5mg/L) + NAA (2.5 mg/L) + KT (0.5mg/L) + supplemented components for dedifferentiation, MS+KT (3.0mg/L) +NAA (0.75mg/L) +IAA (0.5mg/L) +Sorbitol (5.0mg/L) + supplemented components for differentiation. To make the plantlets strong, they were transferred onto the medium of 1/2MS(all Fe-salt)+MET(2.0mg/L). The induction frequency of anther callus was highly related to the density of anthers; The frequency of anther callus induction could be obviously improved in the condition of anther density over certain extent and the optimum density in the bottle of 100ml (about 30ml medium) was 81-120 anthers per bottle. Lower temperature pretreatment with 0-4℃ for 4-14d on panicles before inoculating were beneficial to callus induction. 900 regenerated green plants (374 haploid plants and 526 diploid plants) were obtained in total. By observation of their agronomic characters of 129 DH lines from two hybrids F1 shennong265/haogelao and handao277/ZY-2, their HI generation showed genetic diversity, their H2-H3 generation maintained the relative genetic stability and doesn't exhibit vigor decline. The major lines composed H2 generation anther-cultured plants from lowland rice/upland rice displayed relative uniformity and there were the rich genetic types in the different DH lines.