火炬松体细胞胚胎发生体系的优化  

Optimization of Somatic Embryogenesis for Pinus taeda

作　　者：胡珊 杨春霞[1] 谷振军 杜强[1] 肖平江 李火根[2] HU Shan;YANG Chun-xia;GU Zhen-jun;DU Qiang;XIAO Ping-jiang;LI Huo-gen(Jiangxi Academy of Forestry,Nanchang 330013,Jiangxi,China;Nanjing Forestry University,Co-Innovation Center for Sustainable Forestry in Southern China,Nanjing 210037,Jiangsu,China;Wugongshan Forest Farm of Anfu County,Ji’an 343200,Jiangxi,China)

机构地区：[1]江西省林业科学院,江西南昌330013 [2]南京林业大学南方现代林业协同创新中心,江苏南京210037 [3]安福县武功山林场,江西吉安343200

出　　处：《林业科学研究》2022年第3期9-17,共9页Forest Research

基　　金：江西省重点研发计划项目(20203BBF63022);江西省林业局林业科技创新专项经费(201601)。

摘　　要：[目的]建立和优化火炬松体胚发生技术体系,为火炬松优良基因型体胚繁育及遗传转化体系建立提供技术支撑。[方法]以火炬松优良基因型的未成熟合子胚为材料,从基因型、球果采集期、基本培养基、植物外源激素组合及浓度等方面对胚性愈伤组织诱导条件进行优化;选取增殖效果好、具有胚性胚柄细胞团(ESM)的胚性愈伤组织开展体胚成熟的正交试验,选择发育正常的子叶胚进行萌发,8周后移栽。[结果]基因型、球果采集期、基本培养基和外源激素(PGR)组合及浓度等因子均对胚性愈伤组织诱导有不同程度的影响。火炬松6种基因型中荣山52胚性愈伤组织诱导率最高,达63.33%;处于裂生多胚期的合子胚为外植体的最佳采样期(7月20日左右);6种基本培养基中,在DCR基本培养基上胚性愈伤组织诱导率最高,达40.00%;不同PGR组合及浓度中,以0.5 mg·L^(−1)6-BA+1.0 mg·L^(−1)2,4-D+2.0 mg·L^(−1) NAA+5.0 mg·L^(−1) ABA的胚性愈伤组织诱导率最高,达50.00%。胚性愈伤组织在基本培养基MLP上培养14 d后,转入含有ABA 8.0 mg·L^(−1)、PEG 8000140 g·L^(−1)以及麦芽糖40.0 g·L^(−1)的体胚成熟分化培养基上,体胚发生数平均为411个·g^(−1),体胚萌发率可达93.33%;体胚苗移栽成活率可达86.30%。[结论]本研究优化了火炬松胚性愈伤组织诱导与体胚发生技术体系,体胚诱导率显著提升,为火炬松良种规模化繁育及遗传转化提供技术支撑。[Objective]To provide technical support for the establishment of somatic embryogenesis multiplication and genetic transformation of Pinus taeda,the somatic embryogenesis technology system of P.taeda were established and optimized.[Methods]The immature zygotic embryos with superior genotypes of P.taeda were used.The induction conditions of embryogenic callus were optimized based on the genotype,cone collection period,basic medium,plant hormone combination,and concentration,etc.The embryogenic callus with good proliferation and embryo suspensor mass were selected to carry out the orthogonal test of somatic embryo maturation.In addition,the cotyledon embryos with normal development were selected for germinating.And after 8 weeks,transplanting was carried out.[Results]There were significant effects of genotype,development stage of zygote embryo,basic medium,plant growth regulator combinations,and concentrations on embryogenic callus induction.Among the six genotypes of P.taeda,the induction rate of Rongshan 52 genotype was the highest,63.33%.The best sampling time for explants was the zygotic embryo in the split polyembryonic stage(around July 20).The embryogenic callus induction rate on DCR medium was the highest,34.00%.The induction rate in the PGR combinations and concentrations with 2,4-D 1.0 mg·L^(−1),6-BA 0.5 mg·L^(−1),NAA 2.0 mg·L^(−1) and ABA 5.0 mg·L^(−1) was the highest,50.00%.Embryogenic calli were cultured on MLP for 14 days,and then transferred to somatic embryo induction medium containing ABA 8.0 mg·L^(−1),maltose 40.0 g·L^(−1) and PEG 8000140 g·L^(−1).The average number of somatic embryos was 411·g^(−1),the somatic embryo germination rate was 93.33%,and the survival rate of somatic embryo seedling transplantation was more than 80%.[Conclusion]Somatic embryogenesis of P.taeda was optimized,and the induction rate of somatic embryos was significantly improved.This study provides a preliminary technique for the establishment of large-scale embryogenic transformation of P.taeda.

关 键 词：火炬松 未成熟合子胚 胚性愈伤组织 体细胞胚胎发生 体胚萌发 

分 类 号：S791.255[农业科学—林木遗传育种]