碳氮磷营养对百蕊草悬浮培养细胞生长的影响  

Effects of Carbon,Nitrogen and Phosphorus Nutrition on Growth of Suspension Culture Cells of Thesium chinese Turcz.

作　　者：黄燕芬 王自布 施琴 谢奇 HUANG Yan-fen;WANG Zi-bu;SHI Qin;XIE Qi(School of Biological Sciences,Guizhou Normal University,Guizhou Guiyang 550018,China;Guizhou Characteristic Key Laboratory of Biological Resource Exploitation and Utilization,Guizhou Guiyang 550018,China)

机构地区：[1]贵州师范学院生物科学学院,贵州贵阳550018 [2]贵州省生物资源开发利用特色重点实验室,贵州贵阳550018

出　　处：《西南农业学报》2021年第12期2580-2585,共6页Southwest China Journal of Agricultural Sciences

基　　金：贵州省科技厅自然科学研究基金项目[黔科合基(2018)1117];贵州省科技厅自然科学研究基金项目[黔科合基础(2016)1415];贵州省教育厅自然科学研究计划项目[黔教合KY字(2015)358号];贵州省一流师资团队建设项目[黔教高发(2017)158]。

摘　　要：【目的】建立良好的百蕊草悬浮细胞培养体系,为后续利用基因工程改造百蕊草代谢途径获得高次生代谢产物新品种,实现野生资源的替代奠定基础。【方法】以百蕊草愈伤组织为材料,探寻百蕊草悬浮细胞生长与营养消耗规律的基础上,优化培养液氮和磷的初始投入量及铵态氮和硝态氮的比例(NH_(4)^(+)/NO_(3)^(-))。【结果】在30 d的培养过程中,悬浮细胞生长曲线出现较明显的延滞期(0~6 d)、对数生长期(7~15 d)、稳定生长期(16~24 d)和衰亡期(24 d后);百蕊草悬浮细胞培养28 d时,起始总氮浓度70 mmol/L、磷酸盐浓度1.3 mmol/L情况下,悬浮细胞干物质量均达最大值,低于或高于此两浓度都不利于细胞的健康生长;在此基础上进行悬浮培养细胞对硝态氮、铵态氮的需求探究。结果发现,培养过程中悬浮细胞生长对硝态氮的消耗多于铵态氮,铵态氮的利用消耗先于硝态氮,铵态氮的浓度过高会抑制细胞生长,NH_(4)^(+)/NO_(3)^(-)为20/50最适于细胞生长。【结论】在培养周期内百蕊草悬浮细胞生长曲线呈"S"形;以MS为基本培养基,氮浓度为70 mmol/L,磷酸盐浓度为1.3 mmol/L,NH_(4)^(+)/NO_(3)^(-)为20/50较有利于百蕊草细胞的悬浮培养;培养基总糖、蔗糖、还原糖以及硝态氮、无机磷的消耗均在其对数生长期较快,铵态氮在生长初期消耗快于硝态氮。【Objective】A good suspension cell culture system of Thesium chinese Turcz.is established,so as to lay a foundation for the subsequent use of genetic engineering to transform the metabolic pathway of Thesium chinese Turcz.,obtain new varieties of high-order metabolites,and realize the substitution of wild resources.【Method】On the basis of exploring the law of growth and nutrient consumption of Thesium chinese Turcz.suspension cells,the initial input of liquid nitrogen and phosphorus and the ratio of ammonium nitrogen to nitrate nitrogen(NH_(4)^(+)/NO_(3)^(-))were optimized.【Result】In the 30 days culture process,the growth curve of suspension cells showed an obvious delay period(0-6days),logarithmic growth period(7-15 days)),stable growth period(16-24 days)and decay period(after 24 days).The results showed that the dry matter mass of suspension cells reached the maximum when the initial total nitrogen concentration was 70 mmol/L and the phosphate concentration was 1.3 mmol/L for 28 days,and it was unhealthy for the growth of cells when the nitrogen concentration and phosphate were higher and lower.On this basis,the demand of suspension cultured cells for nitrate nitrogen and ammonium nitrogen were explored.The results showed that the suspension cells during the culture process consumed more nitrate nitrogen than ammonium nitrogen,while the utilization and consumption of ammonium nitrogen precedes that of nitrate nitrogen and too high concentration of ammonium nitrogen would inhibitcell growth.It was most suitable for cell growth when NH_(4)^(+)/NO_(3)^(-)was 20/50.【Conclusion】The growth curve of Thesium chinese Turcz.suspension cells shows‘S’as taking MS as the basic medium,the concentration of nitrogen is 70 mmol/L,the concentration of phosphate is 1.3 mmol/L,and the concentration of NH;/NO;is 20/50,which is more conducive to the suspension culture of Thesium chinese cells.Under these conditions,the consumption of total sugar,sucrose,reducing sugar,nitrate nitrogen and inorganic phosphorus are

关 键 词：百蕊草 基质营养物质 悬浮细胞 

分 类 号：S567.239[农业科学—中草药栽培]