Screening of housekeeping genes in Gelsemium elegans and expression patterns of genes involved in its alkaloid biosynthesis / 生物工程学报

Gelsemium elegans is a traditional Chinese herb of medicinal importance, with indole terpene alkaloids as its main active components. To study the expression of the most suitable housekeeping reference genes in G. elegans, the root bark, stem segments, leaves and inflorescences of four different parts of G. elegans were used as materials in this study. The expression stability of 10 candidate housekeeping reference genes (18S, GAPDH, Actin, TUA, TUB, SAND, EF-1α, UBC, UBQ, and cdc25) was assessed through real-time fluorescence quantitative PCR, GeNorm, NormFinder, BestKeeper, ΔCT, and RefFinder. The results showed that EF-1α was stably expressed in all four parts of G. elegans and was the most suitable housekeeping gene. Based on the coexpression pattern of genome, full-length transcriptome and metabolome, the key candidate targets of 18 related genes (AS, AnPRT, PRAI, IGPS, TSA, TSB, TDC, GES, G8H, 8-HGO, IS, 7-DLS, 7-DLGT, 7-DLH, LAMT, SLS, STR, and SGD) involved in the Gelsemium alkaloid biosynthesis were obtained. The expression of 18 related enzyme genes were analyzed by qRT-PCR using the housekeeping gene EF-1α as a reference. The results showed that these genes' expression and gelsenicine content trends were correlated and were likely to be involved in the biosynthesis of the Gelsemium alkaloid, gelsenicine.

Whole-genome sequencing and analysis of the Chinese herbal plant

Background@# () (2n = 2x = 16) is genus of flowering plants belonging to the Gelsemicaeae family.@*Method@#Here, a high-quality genome assembly using the Oxford Nanopore Technologies (ONT) platform and high-throughput chromosome conformation capture techniques (Hi-C) were used.@*Results@#A total of 56.11 Gb of raw GridION X5 platform ONT reads (6.23 Gb per cell) were generated. After filtering, 53.45 Gb of clean reads were obtained, giving 160 × coverage depth. The genome assemblies 335.13 Mb, close to the 338 Mb estimated by k-mer analysis, was generated with contig N50 of 10.23 Mb. The vast majority (99.2%) of the assembled sequence was anchored onto 8 pseudo-chromosomes. The genome completeness was then evaluated and 1338 of the 1440 conserved genes (92.9%) could be found in the assembly. Genome annotation revealed that 43.16% of the genome is composed of repetitive elements and 23.9% is composed of long terminal repeat elements. We predicted 26,768 protein-coding genes, of which 84.56% were functionally annotated.@*Conclusion@#The genomic sequences of could be a valuable source for comparative genomic analysis in the Gelsemicaeae family and will be useful for understanding the phylogenetic relationships of the indole alkaloid metabolism.

Effect of koumine on the inflammation of RAW264.7 cells induced by LPS / 中国兽医学报

To reveal the anti-inflammation mechanism of koumine,we determined effects of different concentrations of koumine(100,200,400 mg/L) on the secretion of NO,IL-1β,IL-6 and TNF-a by nitrate reductase and ELISA as well as the mRNA of iNOS,IL-1β,IL-6 and TNF-α.Western blot was used to detect iNOS protein expression.The results showed that 100,200,400 mg/L of koumine can significantly inhibit the secretion of NO,IL-1β,IL-6 and TNF-α of RAW264.7 cell (P＜0.01).Koumine can dose-dependently down-regulate the mRNA of iNOS,IL-1β,IL-6 and TNF-α,meanwhile koumine can also significantly inhibit the protein expression of iNOS.The results indicated that koumine may play an anti-inflammation activity by mean of the reduction of NO and mediator of inflammation,down-regulating the mRNA expression of iNOS,IL-1β,IL-6 and TNF-a,decreasing protein expressions of iNOS.