Cloning and functional characterization of pathogenesis-related PR10-1 gene in Panax notoginseng / 药学学报

With homology cloning approaches coupling with RACE (rapid-amplification of cDNA ends) techniques, the full-length coding sequence of pathogenesis-related protein PR10-1 with differential expression was cloned from the total RNA of the root of Panax notoginseng, and its function was explored furtherly. As a result, the longest 465 bp ORF (named as PnPR10-1 with the Accession No. KJ741402 in GenBank) was detected from the cloned sequence with full-length of cDNA of 863 bp. The corresponding peptide encoded consisted of 155 amino acids, contained some domains such as Bet-v-I, and showed high similarity with that from Panax ginseng by analysis of phylogenetic trees created from the alignments. Real-time quantitative PCR showed that the expression of PnPR10-1 gene was constitutive in different tissues of 1-3 year old plant, suggesting that it might be involved in growth, development, and secondary metabolism; yet it was up-regulated significantly with the infection of Fusarium oxysporum in root, suggesting that it might be involved in defense against many diseases including root rot in P. notoginseng.

Cloning and expression analysis of GGPPS gene from Panax notoginseng / 中国中药杂志

According to the transcriptome dataset of Panax notoginseng, the key geranylgeranyl pyrophosphate synthase gene (GGPPS) in terpenoid backbone biosynthesis was selected to be cloned. Using specific primer pairs combining with RACE (rapid amplification of cDNA ends) technique, the full-length cDNA sequence with 1 203 bp, which containing a 1 035 bp open reading frame, was cloned and named as PnGGPPS. The corresponding full-length DNA sequence contained 2 370 bp, consisted of 1 intron and 2 exons. The deduced protein PnGGPPS contained 344 amino acids and shared more than 73% identity with GGPPS from Ricinus communis and Salvia miltiorrhiza. PnGGPPS also had specific Aspartic acid enrichment regions and other conserved domains, which belonged to the Isoprenoid-Biosyn-C1 superfamily. The quantitative real-time PCR showed that PnGGPPS expressed in different tissues of 1, 2, 3 years old root, stem, leaf and 3 years old flower, and the expression level in 3 years old leaf was significant higher than that in other organs, which suggested that it might not only be involved in the regulation of the growth and development, but also be associated with the biosynthesis of chlorophyll and carotenoids, the development of chloroplast, the shade habit and the quality formation of P. notoginseng.