International Journal of Horticulture and Floriculture ISSN 2756-3790 Vol. 9 (1), pp. 001-004, January, 2021. © International Scholars Journals

Full Length Research Paper

Quantitative determination of DNA Methylation in tobacco leaves by HPLC

Feng Jia¹, YunPeng Fu¹, WeiQun Liu²*, Zheng Du¹ and YiDan Zhao²

¹College of Tobacco Science, Henan Agricultural University, Zhengzhou 450002, People’s Republic of China.

²College of Life Sciences, Henan Agricultural University, Zhengzhou, 450002, People’s Republic of China.

Accepted 20 October, 2020

Abstract

A relatively straightforward high performance liquid chromatography (HPLC) method for determination of DNA methylation in flue-cured tobacco leaves was developed and validated. DNA methylation was measured as follows: DNA was hydrolyzed to nucleotide units in perchloric acid. The bases of cytosine and 5-methylcytosine were separated on a reversed-phase C18 column at 30°. The mobile phase used methanol: 5 mmol/L pentane sulfonate (10:90, v/v) and a flow rate of 0.7 mL/min. The absorbance of bases was measured at 273 nm. The results showed that the method was accurate, quick, and cost effective to determine DNA methylation of flue-cured tobacco.

Key words: Tobacco, DNA methylation, 5- Methylcytosine, high performance liquid chromatography (HPLC).