【作者】 孙榕江；

【导师】 李阳春； 梁慧敏； 刘自学；

【作者基本信息】 甘肃农业大学 ， 野生动植物保护与利用， 2006， 硕士

【摘要】 匍匐翦股颖(Agrostis stolonifera L．)是一种多年生冷季型草坪草，具有耐修剪、观赏性高等优点，在世界各国广泛应用，是我国北方地区建植高档草坪的重要草种。 本试验以匍匐翦股颖的成熟种子为外植体，研究了不同匍匐翦股颖种子在愈伤组织诱导、芽的分化以及根的诱导等过程中，不同基本培养基、不同激素浓度及配比等条件下对其再生体系建立的影响，并找出了适宜匍匐翦股颖再生体系建立的最佳条件，得到一套完整的组培再生体系，并对其遗传转化进行了初步研究，为匍匐翦股颖目的基因的导入打下了基础。主要研究结果如下： 1．外植体消毒：匍匐翦股颖的种子在流水中冲洗30min，70％的酒精消毒30s，0.1％升汞消毒12～15min，此种消毒处理效果良好。 2．愈伤组织的诱导：在MS+2，4-D2mg／L+6-BA0.1mg／L+蔗糖30g／L+琼脂7g／L培养基上，暗培养2周，三个品种愈伤组织的诱导率达到最大值，L-93为78.7％，pennA-1 60.8％，pennA-4 59.8％。 3．愈伤组织的继代：在MS+2，4-D2mg／L+6-BA0.1mg／L+蔗糖30g／L+琼脂7g／L愈伤组织继代培养基上，愈伤组织生长快，结构紧实，容易分化，不出现纤毛结构。 4．芽的诱导：诱导匍匐翦股颖愈伤组织芽的分化时，采用MSO+蔗糖15g／L+琼脂7g／L培养基，不添加任何激素，愈伤组织芽的分化率达到85％以上。 5．根的诱导：诱导生根在MSO+IBA0.2mg／L+蔗糖15g／L+琼脂7g／L培养基上进行时，生根率达70％以上。其中L-93可达到88.8％。 6．组培苗的移栽：选取健壮幼苗移栽，并经驯化炼苗15-20天后，大田成活率可达90％以上。 7．确定了对农杆菌LBA4404有明显抑制作用效果的抗生素类型和适宜的浓度：抗菌素采用头孢霉素，最佳抑菌浓度为300mg／L。 8．确定潮霉素抑制植物细胞生长、分化的临界浓度为50mg／L。更多还原

【Abstract】 Agrostis stolonifera L., a widely-grown cool season turf garss, with beautiful colour and the adventage of cut-resistance, is used in planting-lawn in north china and the whole globe.Mature seeds of creeping bentgrass were applied as explant material in all experiments. The effect of different medium, different concentrations of phytohormones on callus inducing , shoots differentiation and roots inducing were studied. The optimum conditions were found, providing a basis for gene transformation. A full regeneration system of Agrostis stolonifera L. was achieved. The main results of this research were listed as following:1. Disinfection of explants: The seeds were washed in water for 30 minutes, before putting in 70% ethanol for 30s. Then the seeds were surface-disinfected for 12～15 minutes in HgCl₂（0.1%）. This kind of treatment can achieve good disinfection effect.2. Callus induction: The effects of different medium and phytohormones on callus induction were studied, which show that MS medium appended with 2mg/L2,4-D , 0.1mg/L 6-BA, 30g/L sucrose and 7g/L agar can give highest rate of callus induction, L-93 was 78.7%, pennA-1 was 60.8%, pennA-4 was 59.8%。3. Callus subculture: MS medium supplemented with 2,4-D2.0 mg/L, 6-BA0.1 mg/L and sucrose 30g/L is the most effective subculture medium.4. The best medium for shoot differentiation is MSO medium supplemented with sucrose 15g/L without phytohormones, and the shoot differentiation frequency is as high as 85%.5. Roots induction: The suggested medium is MSO+IBA0.2mg/L+sucrose 15g/L, which give 70% upwards, and L-93’s roots induction rate is up to 88.8%.6. Tube-plants transfer: The strong tube-plants with more roots were selected and acclimated in green house for 15～20 days, before transferred into the field. The surviving rate is up to 90%.7. Different bacteriophages were studied in controlling agrobacterium LBA4404.更多还原