贮藏方法对紫花苜蓿种子内生根瘤菌传代能力的影响

Effects of storage methods on the transmissibility of endophytic rhizobia from alfalfa seeds

摘要:

为探索贮藏方法对紫花苜蓿种子内生根瘤菌传代能力的影响,将青色荧光蛋白标记根瘤菌gn5f(内源)和12531f(外源)接种于结荚期甘农5号紫花苜蓿。收获的种子,在25 ℃+自然湿度(S1)、25 ℃+硅胶干燥(S2)、4 ℃+冰箱冷藏湿度(S3)和-4 ℃+冰箱冷冻湿度(S4)条件下贮藏3年,然后播种于大田,次代植株苗期进行内生根瘤菌传代能力和传代有效性检测,以无菌水浇灌苜蓿植株后收获的种子为对照。结果表明:标记根瘤菌gn5f和12531f均可通过种子传代到次代植株。S1贮藏条件下,gn5f在次代植株根、茎、叶的定殖数量显著高于其他处理(P＜0.05),次代植株苗期单株结瘤数、单株根瘤重、根瘤等级、固氮酶活性、单株叶片数、株高、根长、地上部分鲜重、地上部分干重、根鲜重、根干重较CK分别增加了37.00%、1 183.54%、47.52%、23 909.07%、38.72%、40.63%、33.82%、68.11%、137.73%、223.70%、600.00%。S2贮藏条件下,12531f在次代植株根部定殖数量最高,为560.63 cfu/g,次代植株苗期单株结瘤数、根瘤直径、根瘤等级、固氮酶活性、单株叶片数、株高、根长、地上部分鲜重、地上部分干重、根鲜重、根干重较CK分别增加了37.90%、21.33%、27.01%、288.21%、41.22%、39.40%、36.04%、72.37%、144.58%、284.05%、872.04%。适宜的贮藏方法有利于提高种子内生根瘤菌的传代能力和传代有效性,但因根瘤菌株种类和来源而异。gn5f内生种子在S1条件下贮藏、12531f内生种子在S2条件下贮藏分别有助于gn5f和12531f传代到次代植株并高效结瘤固氮,促进次代植株生长;且S2条件贮藏的12531f内生种子形成的次代植株结瘤指标和生长指标显著优于S1条件贮藏的gn5f内生种子形成的次代植株。

Abstract:

Seed endophytic rhizobia migration is a new form of sustainable symbiosis system to promote the growth of alfalfa plants.In order to explore the effect of storage methods on the transmissibility of seed endo-phytic rhizobia,seeds were harvested from Medicago sativa cv.Gannong No.5 plants which were inoculated with green fluorescent protein-labeled rhizobia gn5f (endogenous) and 12531f (exogenous).Before sowing,the seeds were stored for 3 years under the following conditions:25 ℃+natural humidity (S1),25 ℃+silica gel (S2),4 ℃+refrigerator humidity (S3) and -4 ℃+refrigerator freezing humidity (S4).The control was the seeds harvested from alfalfa plants irrigated with sterile water.The sustainability and effectiveness of endophytic rhizobia were evaluated at the seedling stage of the subsequent generation.Results showed that gn5f and 12531f can be passed from seeds to the next generation plants.Under S1 condition,the number of gn5fcolonized in the roots,stems and leaves of the next generation plants were significantly higher than that under other treatments (P＜0.05).At the seedling stage of the next generation plants,compared with the control,the nodule number per plant,nodule weight per plant,nodule grade,nitrogenase activity,leaf number,plant height,root length,above ground fresh weight,above ground dry weight,root fresh weight,root dry weight were increased by 37%,1183.54%,47.52%,23909.07%,38.72%,40.63%,33.82%,68.11%,137.73%,223.70% and 600.00%,respectively.Under S2 condition,12531f had the highest colonization number in the roots of the next generation plants (560.63 cfu/g).The nodule number per plant,nodule diameter,nodule grade,nitrogenase activity,leaf number per plant,plant height,root length,above ground fresh weight,above ground dry weight,root fresh weight,root dry weight were higher than those of control and increased by 37.90%,21.33%,27.01%,288.21%,41.22%,39.40%,36.04%,72.37%,144.58%,284.05%,872.04%,respectively.Appropriate storage methods contributed to the improvement of sustainability and effectiveness of seed endophytic rhizobia,but it varied with the species and source of rhizobia.Storage of gn5f-colonized seeds under S1 condition and 12531f-colonized seeds under S2 condition demonstrated higher transmissibility to the subsequent generation and better promotion of plant growth by efficient nodulation and nitrogen fixation.The next generation plants of 12531f-colonized seeds stored under S2 condition presented higher nodulation and growth parameters than that of gn5f-colonized seeds stored under S1 condition.