摘  要从青海盐湖水中分离纯化得到4种菌株，标记为QH1、QH3、QH4、QH5。菌株的形态都呈现杆状，QH1、QH3、QH5为革兰氏阳性菌，QH4为革兰氏阴性菌。提取嗜盐菌株基因组总DNA序列，使用16S rRNA基因序列的通用引物，对总DNA序列进行PCR扩增，对PCR扩增产物进行测序。使用ContigExpress软件将测序后结果进行拼接，得到1480bp完整连续的QH1 16S rRNA序列。使用EzTaxon数据库和MEGA7.0软件对QH1进行分类比较并构建系统进化树，初步确定QH1属于枝芽孢杆菌属（Virgibacillus）。进一步研究QH1的生长特性，得出该菌0-34小时为生长延滞期，34-42小时为生长对数期，42小时以后进入生长稳定期。最后用不同浓度梯度的KCl溶液对QH1进行胁迫，基于流式细胞分析技术，结果得出QH1生长环境适宜的KCl浓度为0.030 mol/L。

该论文有图7幅，表4个，参考文献14篇。69930

毕业论文关键词：嗜盐细菌  16S rRNA基因序列  系统进化  胁迫  流式细胞分析术

Study on Salt-tolerance Characteristic of Halophiles under KCl Stress

Abstract 4 strains were isolated and purified from the water of Qinghai Lake, which were labeled as QH1, QH3, QH4 and QH5. The strains were in the form of rods. QH1, QH3 and QH5 were gram positive bacteria and QH4 was gram negative bacteria. The total genomic DNA sequences of the strains were extracted.The common primers of 16S rRNA gene were used to amplify the total gene sequence by PCR and the PCR amplification products were sequenced. ContigExpress software was used to splice the results of sequencing and the 1480bp QH1 16S rRNA sequence was obtained. EzTaxon database and MEGA7.0 software were used to classify the QH1 and to construct phylogenetic trees. The QH1 is determined preliminarily to belong to  Virgibacillus. To study further on growth characteristics of QH1 and then draw the conclusion: 0-34 hours is the lag phase of QH1’s growth;34-42 hours is the phase of logarithmic growth;42 hours later is the stationary phase of growth. Finally, based on flow cytometry, QH1 was subjected to KCl stress with different concentration gradient.The results showed that the suitable KCl concentration for QH1’s growth was 0.030 mol/L.

Key Words: Halophiles  16S rRNA gene sequences  stress  flow cytometry  phylogenetic trees

目  录

摘要Ⅰ

Abstract-Ⅱ

目录Ⅲ

图清单-Ⅳ

表清单-Ⅳ

1 绪论-5

2 材料与方法5

2.1材料-5

2.2方法-6

3 结论与分析9

3.1嗜盐菌菌落形态及菌体特性-10

3.2嗜盐菌株PCR产物电泳检测分析-10

3.3嗜盐菌株测序结果与分析10

3.4嗜盐菌株系统进化树分析10

3.5嗜盐菌株QH1的生长曲线测定-11

3.6嗜盐细菌QH1受KCl溶液胁迫最适处理时长的研究11

3.7嗜盐细菌QH1最适生长KCl溶液浓度的研究-12

4 讨论14

参考文献15

致谢16

图清单

图序号 图名称 页码

图3-1 16S rRNA PCR产物电泳结果 10

图3-2 基于16S rRNA序列的菌株QH1与邻近菌株构建的分子发育树 11

图3-3 菌株QH1的生长曲线 11

图3-4 菌株QH1在不同胁迫时长后的死活细胞比例 12

图3-5 菌株QH1受KCl盐梯度胁迫3小时后的死活细胞比例（1）