Cloning And Sequence Analysis Of Chalcone Synthase, Flavonol Synthase And Chalcone Isomerase Gene From Lonicera Japonica

In the present study, the full length cDNA of Chalcone synthase (chs), flavonol synthase (fls) and Chalcone Isomerase (chi) from Lonicera japonica leaves were cloned with RT-PCR and RACE, which set up a foundation for future study of the relationship between these genes and Flavonoids Biosynthesis in molecular level, for revealing of the molecular mechanism of Flavonoids Biosynthesis and Flavonoids exploitation in Lonicera japonica Leaf. The results were as follows:1Cloning of the full length cDNA of Chalcone synthase (chs) from Lonicera japonica leavesThe full length cDNA of Lonicera japonica leaves chs is1467bp. It contains an ORF(1167bp) encoding389amino acids(Mw=42.7kD, pI=7.49), a5'untranslated region of118bp and a3'flanking sequence of179bp. Compared with chs gene of others plants by Blast software, Lonicera japonica leaves chs cDNA respectively had83%,82%,81%identity with that of Populus trichocarpa, Nelumbo nucifera and Malus hybrid chs genes. The comparison among the putative amino acid sequence with Populus trichocarpa CHS showed that identities were93%. This cDNA had been registered in GeneBank with accession number JQ627646.2Cloning of the full length cDNA of flavonol synthase(fls) from Lonicera japonica leavesA pair of primers was designed according to the conserved cDNA sequence of flavonol synthase (fls) gene of other species and partial cDNA sequence of fls was cloned from Fonicera japonica leaves using RT-PCR and RACE. The results indicated that the full length cDNA sequence of fls was1248bp and it included an open reading frame (ORF) which encoded333amino acids (Mw=37.7kD, pI=5.45), a5' untranslated region of38bp and a3'flanking sequence of208bp. This cDNA had been registered in GeneBank with accession number JQ627647. Sequence analysis showed that the homologies of fls cDNA from Fonicera japonica with Nicotiana tabacum, Solanum tuberosum and Platycodon grandiflorus were86%,83%,80%and among the others were about80%,which revealed that the fls gene kept highly conserved among different species.3Cloning of the full length cDNA of Chalcone Isomerase (chi) from Lonicera japonica leavesThe full length cDNA of Lonicera japonica leaves chi is725bp. It contains an ORF(630bp) encoding210amino acids (Mw=22.5kD, pI=6.48), a5' untranslated region of40bp and a3'flanking sequence of55bp. Compared with chi gene of others plants by Blast software, Lonicera japonica leaves chi cDNA respectively had80%,79%,76%identity with that of Fragaria ananassa, Rhododendron simsii Planch and Vitis vinifera chi genes. This cDNA had been registered in GeneBank with accession number JQ650259.Above all, chs, fls and chi genes might play an important role in flavonoids synthesis of the Lonicera japonica leaves. The cloning of these three genes will contribute to further development and utilization of the flavonoids in Lonicera japonica leaves.