烤烟烟碱含量标记基因NtMPO1和NtARF6的鉴定OA北大核心CSTPCD

In order to identify the effective nicotine content marker genes,the nicotine contents of 89 flue-cured tobacco varieties were determined,the high-,medium-and low-nicotine flue-cured tobacco varieties were screened,the relative expression levels of a total of 25 genes related with nicotine synthesis,transport,regulation in high-,medium-and low-nicotine flue-cured tobacco varieties were determined by real-time quantitative PCR(qRT-PCR),the correlation between relative expression levels of the above genes and nicotine content was analyzed,and candidate marker genes for nicotine content were screened.The leaves of tobacco treated with 2.5、5.0 7.5 mmol/L NaNO3,upper,middle and lower parts of leaves and the leaves before and after topping(10 days)were used to determine the nicotine content and the relative expression levels of candidate marker genes,so as to verify the effectiveness of nicotine content candidate marker genes.The results showed that 4 high-,4 medium-and 4 low-nicotine flue-cured tobacco varieties were screened from 89 flue-cured tobacco varieties,and the nicotine content of these flue-cured tobacco varieties was significantly negatively correlated with the relative expression levels of NtMPO1 gene and significantly positively correlated with the relative expression levels of NtARF6 gene,which could be used as candidate genes for nicotine content marker genes.The results of the validation test of nicotine content candidate marker genes showed that compared with the normal nitrogen treatment,the nicotine content of leaves in the high nitrogen treatment significantly increased,the relative expression level of NtMPO1 gene significantly decreased,and the relative expression level of NtARF6 gene significantly increased;the nicotine content of the low nitrogen treatment significantly decreased,the relative expression level of NtMPO1 gene significantly increased,and the relative expression level of NtARF6 gene significantly decreased.The nicotine content of the upper leaves was significantly higher than that of the middle leaves,and the nicotine content of the middle leaves was significantly higher than that of the lower leaves;the relative expression level of NtMPO1 gene in the upper leaves was significantly lower than that in the middle leaves,and the relative expression level of NtMPO1 gene in the middle leaves was significantly lower than that in the lower leaves;the relative expression level of NtARF6 gene in the upper leaves was significantly higher than that in the middle leaves,and the relative expression level of NtARF6 gene in the middle leaves was significantly higher than that in the lower leaves.Compared with before topping,the nicotine content of leaves significantly increased after topping,the relative expression level of NtMPO1 gene significantly decreased,and the relative expression level of NtARF6 gene significantly increased.In conclusion,the NtMPO1 and NtARF6 genes can be used as effective markers of nicotine content.