Fecha de publicación: 09/11/2016 Fuente: WIPO "tobacco agriculture" The invention discloses a cloning method of a gene NtFLS2 with a function of increasing rutin content of tobacco leaves. The cloning method includes steps: A, using TRIzol for extracting total RNA of tobacco leaves and flowers, and performing reverse transcription to obtain a first strand of cDNA; B, taking the first strand of cDNA as a template, and adopting a primer for PCR (polymerase chain reaction) amplification; C, connecting a target gene segment to a pTOPO carrier to carry out sequencing. The invention further discloses application of the gene NtFLS2 with a function of increasing the rutin content of the tobacco leaves. By cloning of the gene NtFLS2, development and utilization of resources of the gene NtFLS2 lay a foundation for industrial extraction of rutin from the tobacco leaves. According to functional verification of transgenic tobacco plants, the cloned gene NtFLS2 has the function of increasing the rutin content of the tobacco leaves.