Abstract: Expression of cytoplasmic membrane ATPase gene in plants can cause cells to release H⁺ to the external environment, thus form the membrane inside and outside electrical chemical formula of the gradient, promote all sorts of ion transport, various stress resistance to the outside world. In order to study the regulatory mechanism of ATPase in tobacco and its related biological functions, this article adopted homologous cloning method to clone a ATPase4 homologous gene from common tobacco K326 and construct ATPase4-pcambia1300 overexpression vector. The bioinformatics software was used to predict the hydrophobic, physical and chemical properties, protein structure, signal peptide, phosphate site and origin of ATPase4. The homology analysis showed that the modified gene had 99% homology with plasma membrane ATPase4 of Nicotiana tomentosiformis, so it was named as plasma membrane ATPase4 gene. Fluorescence quantitative real time PCR (qRT-PCR) was used to determine the relative expression pattern in different tissues under different stresses. The results showed that the length of gene sequence was 2 982 bp, encoding 963 amino acid residues, the gene had the highest expression level in the roots, and it could respond quickly to low potassium, high salt, PEG and cold stress. It was shown that ATPase4 played an important regulatory role in tobacco stress.

Key words: tobacco, ATPase, cloning, expression