Isolation, Identification, And Genetic Diversity Of Cold-tolerant Endophytic Bacteria From Different Kind Of Citrus

Endogenous bacteria belong to a kind of various and diverse microbial community, and their various kinds of metabolic products contain many disease-resistant, insecticidal, and bacteriostatic substances, which can provied us a novel and effective way to find new disease-resistant medicine. In addition, a substantial numbers of citrus endogenous bacteria play an important role in resisting against cold injury and diseasese and maintaining the quality of citrus fruit.Eight different kinds of citrus that had been dealt with surface sterilization and cold treatment were adopted in the experiments. The eight endogenous bacteria strains were purified, isolated, and acquired on beef extract peptone medium and LB medium using dilution spread plate method and lineation separation method.In this study, based on the conventional classification identification methods, shape of microbial community, their growth condition, and physiological and biochemical characters were monitored and determined by various kinds of experiments such as gram staining, spore staining, and activity experiments. Furthermore, through using the molecular biology methods, the class of different bacteria strains was primarily identified. Our results showed, the GJ1, GJ2, GJ3, GJ7, which were the endophyte of Penggan, Jicheng, Shatang and Putaoyou respectively, belong to the class of Bacillus. Besides, GJ4, GJ5, which were the endophyte of Jinqian orange and Shatianyou respectively, belong to Brevibacillus class. Furthermore, GJ6, GJ8, which were the endophyte of Xuecheng and Bintangan, belong to Paenibacillus class.This study has performed the analysis of16S rDNA sequences of eight citrus endogenous bacteria, and cluster analysis of the16S rDNA sequences by using MEGA4.0software. And the results revealed that the strain GJ1, GJ2, GJ3, and GJ7shared very high similarity, and therefore belonging to the same class. Similarly, the strain GJ4and GJ5could be classified into the same category. Also, GJ6and GJ8belong to the same class.This study provides us with some valuable insights into how to develop cold-tolerant endophytic bacteria and to conduct cold tolerance research of citrus.