Huanan elephant grass (Pennisetum purpureum cv.Huanan) is widely used as forage in South China.It is also a highly yielding energy crop for ethanol production.However,the lignin in lignocellulose inhibits the activity of bioconversion.In order to explore the lignin regulation network,we isolated 4-coumarate CoA ligase (4CL) from P.purpureum cv.Huanan.A 1 943 bp cDNA sequence of Pp4CL gene was cloned by homology-based RT-PCR.Pp4CL has an open reading frame of 1 662 bp and encodes a 553 amino acid protein with a predicted molecular mass of 59.57 kD and a isoelectric point of 5.2.Pp4CL protein is a hydrophobin protein,and belongs to AFD class I superfamily as it contains an AMP binding domain.In phylogenetic tree,Pp4CL was clustered with At4CL1 and Os4CL1.Two conserved motifs,SSGTGLPKGV and GEICIRG,were found in Pp4CL,indicating that it was a typical plant 4CL.The pGEX-4CL prokaryotic expression vector was constructed and GST-tagged Pp4CL fusion protein with the molecular mass of 88 kD was obtained.This result laid foundation for the western blot and enzymology analysis.The overexpression vector named pBA-4CL was constructed,and three positive transgenic tobacco lines were obtained by leaf disc method.In petiole,acetyl bromide lignin content was greater by approximately 10% in OX-9,16.2% in OX-7,94.6% in OX-4 compared with WT plants.And in base stem,the three transgenic plants had 0.9%,4.0% and 13.5% more acetyl bromide lignin content than CK plant,respectively.In conclusion,overexpression of Pp4CL in tobacco leaded to increasing of lignin content,thus,Pp4CL was related to lignin biosynthesis.This research provides scientific basis for lignin improvement in elephant grass and molecular breeding in forage grass.