马尾松优良种源高效组培育苗技术体系构建

【目的】马尾松是我国南方生态建设和造林用材的主要树种,受种苗良种化限制,人工林生产力水平整体不高,产业发展缓慢。因此,构建高效马尾松优良种质繁育技术体系对推动良种推广利用,加快产业发展,提升行业竞争力很有必要。【方法】以马尾松骨干育种资源桐棉种源为研究对象,未成熟球果合子胚为外植体,通过体胚发生技术获取成熟胚状体作为本试验供试材料,针对马尾松体胚萌发率低、芽苗活性差、生根成苗困难等技术瓶颈,系统分析了活性炭(AC)、基本培养基、植物激素对体胚萌发及萌发体胚芽苗复壮与不定根诱导的影响。【结果】(1)AC能显著提升成熟胚状体萌发培养效果,但过高浓度AC会导致培养效果减弱,其中以0.83 mol/L AC效果最佳。在附加AC基础上,利用高N、低NH4+/NO3-比,K、Ca量适中的1/2MMS基本培养基能进一步提升萌发效果,萌发率可达94.1%。(2)0.42 mol/L AC能有效促进萌发体胚芽苗伸长,在4μmol/L TDZ作用下,腋芽诱导效果好,有效芽增殖系数5.6/35 d,芽高9.2 cm/50 d。(3)经1.2μmol/L NAA+2μmol/L PBZ处理60 d,单芽生根率达94.3%、根条数6.4,移栽3个月后成活率为95.8%。【结论】本研究首次通过体胚发生与器官发生途径相结合的技术构建了高效组培繁育体系,可用于马尾松优良种质的快速繁殖以及遗传转化方面的研究,为马尾松良种产业化以及进一步开展基因工程分子育种方面的研究奠定了坚实基础。

Establishment of an effective protocol for cultivation of tissue cultured seedlings in Pinus massoniana superior provenance

Objective] Pinus massoniana is a main tree species for ecological construction and timber in southern areas of China. However, the productivity of plantation is generally low resulting from the limitation of improved varieties, which leads to the slow development of industrialization in P. massoniana.It is necessary to develop an effective propagation system of elite germplasm for P. massoniana in order to promote the use of improved varieties, accelerate the industrial development, and enhance the competitiveness of industry. Method] In this study, ‘Tongmiansong’(TM), the backbone of P. massoniana breeding resources was used as the research object, zygotic embryos excised from immature cones were applied to explants, and mature somatic embryos(SE) obtained via somatic embryogenesis were used for testing materials. Concerning the technical bottlenecks of P. massoniana tissue culture, including low germination rate, poor shoot growth, and recalcitrance to rooting, effects of active charcoal(AC), basal media, and plant hormones on SE germination as well as reinvigoration and adventitious rooting of shoots were investigated in the present study. Result](1) AC significantly improved germination of TM mature SE, while a high level of AC was able to weaken their germination effects, and the best effect was observed at the 0.83 mol/L AC treatment. Based on the application of AC in the medium, the basal medium composed of high N, low ratio of NH4+/NO3-, and moderate K and Ca furtherly enhanced the germination of SE,reaching 94.1% of germinating rate.(2) 0.42 mol/L AC effectively promoted the elongation of germinated SE. Under the treatment of 4 μmol/L TDZ, induction of axillary buds was better, achieving effective bud proliferation coefficient of 5.6/35 d, and shoot height of 9.2 cm/50 d.(3) After 60 days of 1.2 μmol/L NAA+2 μmol/L PBZ application in the rooting medium, rooting rate was 94.3%, root number was 6.4, and survival rate was 95.8% after 3-month transplanting. Conclusion] The effective breeding system by tissue culture for TM was firstly established via a combined approach of somatic embryogenesis and organogeneis in this study, which would be used for the rapid propagation of elite germplasm for P. massoniana as well as for the research on genetic transformation, providing solid foundation for industrialization of improved varieties and molecular breeding in P. massoniana.