Chloroplast mini-barcodes combined with high resolution melting analysis to identify herbal medicine Difengpi (Illicium difengpi).

Difengpi, derived from the air-dried stem bark of Illicium difengpi and enlisted in the Chinese Pharmacopoeia for its therapeutic effect against common ailments, confronts challenges due to dwindling wild resources and intentional substitution with potentially harmful botanical relatives. The imperative need to authenticate this herbal remedy has led to the development of robust methods. Here, we integrated chloroplast mini-barcoding and high resolution melting (HRM) analysis to distinguish Difengpi from the reported adulterants. We assembled the complete chloroplast (cp) genomes of I. difengpi and substituted close relatives I. jiadifengpi and I. majus, and conducted an in-depth comparative analysis to screen divergent regions for exploiting as DNA mini-barcodes. Despite the conservativeness characterizing the whole cp genomes among these Illicium species, we identified some highly variable regions with promising potential as molecular markers for species identification. Subsequently, we designed DNA mini-barcodes and subjected them to HRM analysis to assess their efficacy in species discrimination. Melting profiles unveiled that mini-barcodes designed from four divergence regions trnL-trnF, trnF-ndhJ, ycf1-ndhF and rpl32-trnL exhibited substantial discriminatory power, distinctly differentiated I. difengpi from I. jiadifengpi, I. majus and I. verum. We tested ten commercially available Difengpi products from online stores and local traditional markets using these four mini-barcodes. All ten samples clustered closely with the reference I. difengpi with genotype confidence higher than 93 %, indicating the presence of the claimed species in these samples, sans any reported toxic adulterants. Consequently, we simulated Difengpi mimic samples utilizing I. jiadifengpi, I. majus and I. verum and subjected them to evaluate the practicability of these mini-barcodes. The outcomes confirmed the precision of the four mini-barcodes in accurately discerning the mimic samples. In conclusion, integrating taxon-specific DNA mini-barcodes with HRM analysis is an efficient strategy for the authentication of species identity within commercial herbal products.

The complete chloroplast genome of Ficus pumila, a functional plant in East Asia.

Ficus pumila L. is a climbing fig commonly used as an ornamental plant. In this study, we sequenced and assembled the complete chloroplast genome of F. pumila. The complete chloroplast genome of F. pumila is 160,248 bp in length which includes a pair of inverted repeats (IRs) of 25,871 bp separated by a large single-copy (LSC) region of 88,405 bp and a small single-copy (SSC) region of 20,101 bp. The overall guanine-cytosine (GC) content of F. pumila cp genome is 35.98%, while the corresponding values of LSC, SSC, and IR sequences are 33.65, 29.05, and 42.65%, respectively. The phylogenetic tree was shown to be consistent with the traditional morphology-based taxonomy of Moraceae. Five plants from the genus Ficus formed a well-supported monophyletic clade with 100% bootstrap value, and F. pumila is closely related to F. hirta, F. carica, and F. racemosa, with a support value of 97%. The complete chloroplast of F. pumila contributes to the growing number of chloroplast genomes for phylogenetic and evolutionary studies in Moraceae.

The complete chloroplast genome sequence of Habenaria dentata (Orchidaceae).

Habenaria dentata is a rare species with high ornamental value in China. In this study, we report the complete chloroplast (cp) genome of H. dentata using the Illumina sequencing data. The total genome of H. dentata is 153,682 bp in length and the GC content is 36.62%, with a pair of inverted repeats (IRs) regions of 26,339 bp each, a large single-copy (LSC) region of 83,963 bp and a small single-copy (SSC) region of 17,041 bp. The cp genome encoded 133 genes, including 87 protein-coding genes (PCG), eight rRNA genes, and 38 tRNA genes. The maximum-likelihood phylogenetic analysis based on 12 cp genomes showed that H. dentata was sister to Habenaria chejuensis and Habenaria ciliolaris. This work will be valuable for genetic and phylogenetic studies on H. dentata.

Discovery and Evaluation of Pyrazolo[3,4-d]pyridazinone as a Potent and Orally Active Irreversible BTK Inhibitor.

The identification and lead optimization of a series of pyrazolo[3,4-d]pyridazinone derivatives are described as a novel class of potent irreversible BTK inhibitors, resulting in the discovery of compound 8. Compound 8 exhibited high potency against BTK kinase and acceptable PK profile. Furthermore, compound 8 demonstrated significant in vivo efficacy in a mouse-collagen-induced arthritis (CIA) model.