Scutellaria polysaccharide mediates the immunity and antioxidant capacity of giant freshwater prawn (Macrobrachium rosenbergii).

The giant freshwater prawn (Macrobrachium rosenbergii) is a commercially valuable freshwater crustacean species that frequently appears a death affected by various diseases, resulting in substantial economic losses. Improving the survival rate of M. rosenbergii is a hot and essential issue for feeding the prawns. Scutellaria polysaccharide (SPS) extracted from Scutellaria baicalensis (a Chinese medicinal herb) is conducive to the survival rate of organisms by enhancing immunity and antioxidant ability. In this study, M. rosenbergii was fed 50, 100, and 150 mg/kg of SPS. The immunity and antioxidant capacity of M. rosenbergii were tested by mRNA levels and enzyme activities of related genes. The mRNA expressions of NF-κB, Toll-R, and proPO (participating in the immune response) in the heart, muscle, and hepatopancreas were decreased after four weeks of SPS feeding (P < 0.05). This indicated that long-term feeding of SPS could regulate the immune responses of M. rosenbergii tissues. The activity levels of antioxidant biomarkers, alkaline phosphatase (AKP), and acid phosphatase (ACP) had significant increases in hemocytes (P < 0.05). Moreover, catalase (CAT) activities in the muscle and hepatopancreas, as well as superoxide dismutase (SOD) activities in all tissues, significantly decreased after four weeks of culture (P < 0.05). The results demonstrated that long-term feeding of SPS could improve the antioxidant capacity of M. rosenbergii. In summary, SPS was conducive to regulating the immune capacity and enhancing the antioxidant capacity of M. rosenbergii. These results provide a theoretical basis for supporting SPS addition to the feed of M. rosenbergii.

The genomes of medicinal skullcaps reveal the polyphyletic origins of clerodane diterpene biosynthesis in the family Lamiaceae.

The presence of anticancer clerodane diterpenoids is a chemotaxonomic marker for the traditional Chinese medicinal plant Scutellaria barbata, although the molecular mechanisms behind clerodane biosynthesis are unknown. Here, we report a high-quality assembly of the 414.98 Mb genome of S. barbata into 13 pseudochromosomes. Using phylogenomic and biochemical data, we mapped the plastidial metabolism of kaurene (gibberellins), abietane, and clerodane diterpenes in three species of the family Lamiaceae (Scutellaria barbata, Scutellaria baicalensis, and Salvia splendens), facilitating the identification of genes involved in the biosynthesis of the clerodanes, kolavenol, and isokolavenol. We show that clerodane biosynthesis evolved through recruitment and neofunctionalization of genes from gibberellin and abietane metabolism. Despite the assumed monophyletic origin of clerodane biosynthesis, which is widespread in species of the Lamiaceae, our data show distinct evolutionary lineages and suggest polyphyletic origins of clerodane biosynthesis in the family Lamiaceae. Our study not only provides significant insights into the evolution of clerodane biosynthetic pathways in the mint family, Lamiaceae, but also will facilitate the production of anticancer clerodanes through future metabolic engineering efforts.

De Novo Assembly and Species-Specific Marker Development as a Useful Tool for the Identification of Scutellaria L. Species.

Scutellaria L. (family Lamiaceae) includes approximately 470 species found in most parts of the world and is commonly known as skullcaps. Scutellaria L. is a medicinal herb used as a folk remedy in Korea and East Asia, but it is difficult to identify and classify various subspecies by morphological methods. Since Scutellaria L. has not been studied genetically, to expand the knowledge of species in the genus Scutellaria L., de novo whole-genome assembly was performed in Scutellaria indica var. tsusimensis (H. Hara) Ohwi using the Illumina sequencing platform. We aimed to develop a molecular method that could be used to classify S.indica var. tsusimensis (H. Hara) Ohwi, S. indica L. and three other Scutellaria L. species. The assembly results for S.indica var. tsusimensis (H. Hara) Ohwi revealed a genome size of 318,741,328 bp and a scaffold N50 of 78,430. The assembly contained 92.08% of the conserved BUSCO core gene set and was estimated to cover 94.65% of the genome. The obtained genes were compared with previously registered Scutellaria nucleotide sequences and similar regions using the NCBI BLAST service, and a total of 279 similar nucleotide sequences were detected. By selecting the 279 similar nucleotide sequences and nine chloroplast DNA barcode genes, primers were prepared so that the size of the PCR product was 100 to 1000 bp. As a result, a species-specific primer set capable of distinguishing five species of Scutellaria L. was developed.

Comparative Genome Analysis of Scutellaria baicalensis and Scutellaria barbata Reveals the Evolution of Active Flavonoid Biosynthesis.

Scutellaria baicalensis (S. baicalensis) and Scutellaria barbata (S. barbata) are common medicinal plants of the Lamiaceae family. Both produce specific flavonoid compounds, including baicalein, scutellarein, norwogonin, and wogonin, as well as their glycosides, which exhibit antioxidant and antitumor activities. Here, we report chromosome-level genome assemblies of S. baicalensis and S. barbata with quantitative chromosomal variation (2n = 18 and 2n = 26, respectively). The divergence of S. baicalensis and S. barbata occurred far earlier than previously reported, and a whole-genome duplication (WGD) event was identified. The insertion of long terminal repeat elements after speciation might be responsible for the observed chromosomal expansion and rearrangement. Comparative genome analysis of the congeneric species revealed the species-specific evolution of chrysin and apigenin biosynthetic genes, such as the S. baicalensis-specific tandem duplication of genes encoding phenylalanine ammonia lyase and chalcone synthase, and the S. barbata-specific duplication of genes encoding 4-CoA ligase. In addition, the paralogous duplication, colinearity, and expression diversity of CYP82D subfamily members revealed the functional divergence of genes encoding flavone hydroxylase between S. baicalensis and S. barbata. Analyzing these Scutellaria genomes reveals the common and species-specific evolution of flavone biosynthetic genes. Thus, these findings would facilitate the development of molecular breeding and studies of biosynthesis and regulation of bioactive compounds.

Enhanced flavonoid production in hairy root cultures of Scutellaria bornmuelleri by elicitor induced over-expression of MYB7 and FNSП2 genes.

For the purpose of the current study, hairy root induction in S. bornmuelleri, which is an important medicinal plant, was examined using a particular protocol. Accordingly, some factors such as four strain types of Agrobacterium rhizogenes (A4, A13, MSU440 and ATCC15834), three different explants, namely stem, petiole and leaf, two co-cultivation media, i.e. full and half-MS were studied. Besides, two inoculation methods including injection and immersion as well as three inoculation times (5, 7 and 10 min) were closely taken into account. Utilizing injection method by MSU440 strain, hairy root induction took place in stem explants, and a remarkable increase in transformation frequency (100%) was observed in half-strength MS medium. Methyl jasmonate (MeJA, 100 µM), methyl-b-cyclodextrin (b-CD, 0.7, 7 and 14 mM) and Chitosan (Chi, 50, 100 and 200 mg/l) were used either individually or in a combined way to elicitation. Based on the HPLC results, production of chrysin, wogonin and baicalein increased 9.15, 10.56 and 13.25 times after elicitation of hairy roots by MeJA + Chi. In addition, transcripts of FNSП2 and MYB7, two important genes involved in the flavonoid biosynthesis pathway, were studies. By applying Chi and MeJA + Chi elicitor, the expression of both genes increased noticeably. It can be concluded that the mentioned hairy root culture system of S. bornmuelleri can be an alternative to flavonoids production. Moreover, there is a direct and positive relationship between the expression of FNSП2 and MYB7 genes as well as the level of three flavonoids.

Transcriptomic Insight into Terpenoid Biosynthesis and Functional Characterization of Three Diterpene Synthases in Scutellaria barbata.

Scutellaria barbata (Lamiaceae) is an important medicinal herb widely used in China, Korea, India, and other Asian countries. Neo-clerodane diterpenoids are the largest known group of Scutellaria diterpenoids and show promising cytotoxic activity against several cancer cell lines. Here, Illumina-based deep transcriptome analysis of flowers, the aerial parts (leaf and stem), and roots of S. barbata was used to explore terpenoid-related genes. In total, 121,958,564 clean RNA-sequence reads were assembled into 88,980 transcripts, with an average length of 1370 nt and N50 length of 2144 nt, indicating high assembly quality. We identified nearly all known terpenoid-related genes (33 genes) involved in biosynthesis of the terpenoid backbone and 14 terpene synthase genes which generate skeletons for different terpenoids. Three full length diterpene synthase genes were functionally identified using an in vitro assay. SbTPS8 and SbTPS9 were identified as normal-CPP and ent-CPP synthase, respectively. SbTPS12 reacts with SbTPS8 to produce miltiradiene. Furthermore, SbTPS12 was proven to be a less promiscuous class I diterpene synthase. These results give a comprehensive understanding of the terpenoid biosynthesis in S. barbata and provide useful information for enhancing the production of bioactive neo-clerodane diterpenoids through genetic engineering.

An Untapped Resource in the Spotlight of Medicinal Biotechnology: The Genus Scutellaria.

BACKGROUND: This review is intended to draw the attention of pharmaceutical and biotechnological communities to the untapped potential of the Scutellaria genus. Skullcaps, as they are more widely known, are found in one of the oldest materia medica in the world, that of ancient Chinese pharmacology, and their numerous wide range of medicinal bioactivities have been studied both in vivo and in vitro. For thousands of years, chemical compounds from the Scutellaria species have been safely used as antitumor, antibacterial, antiviral, anti-inflammatory, antioxidant or hepatoprotective factors. OBJECTIVE: As these effects are well known, reflected in the presence of Scutellaria plants in national pharmacopoeias, it is clear that the plant has yet enormous unexploited potential. The European pharmacological market has turned to the resources of Scutellaria only in the last two decades, and although the construction and clinical processing of a new drug is a long process, the general impression is that very few medical products in pharmacies have been inspired by the phytochemistry of skullcaps. CONCLUSION: This paper presents the current state of knowledge on the wealth of Scutellaria chemical compounds with treatment applications, its tissue culture and biotechnological achievements, especially in the context of the production of secondary metabolites.

[Karyotype and evolutionary trend analysis of Scutellaria species in Jinyun mountain, Chongqing].

Traditional squash method was used to analyze chromosome number and karyotypes of four Scutellaria species in Chongqing Jinyun Mountain Natural Reserve： Scutellaria tsinyunensis, S.yunnanensis, S.franchetiana and S.indica.The result showed that the chromosome numbers were 26 except for S.franchetiana, which had 24 chromosomes.These species were all diploid with metacentric and submetacentric chromosomes.Their karyotypes were symmetrical and primitive.The karyotype formula of S.tsinyunensis is 2n=2x=26=24m+2sm, 1B type, As.k=55.28%; the karyotype formula of S.yunnanensis var.salicifolia is 2n=2x=26=26m, 1B type, As.k=56.11%; the karyotype formula of S.franchetiana is 2n=2x=24=20m+4sm, 2B type, As.k=58.50%; the karyotype formula of S.indica is 2n=2x=24=20m+4sm, 2B type, As.k=58.41%.The results were compared with the reported data of S.baicalensis and S.alaschanica.S.alaschanica is expected to be the most advanced one whereas S.tsinyunensis, and S.yunnanensis var. salicifolia primitive.These results are expected to provide some references to the origin and differentiation of genus Scutellaria.

[Induction and culture of hairy roots in Scutellaria viscidula and its baicalin production].

OBJECTIVE: To establish a culturing system of hairy roots of Scutellaria viscidula, and study the hairy roots growth and biosynthesis of flavonoid in the culturing system. METHOD: Hairy roots of S. viscidula were obtained from infected stem explants after infection with the disarmed Agrobacterium tumefaciens strain C58C1, elite strains were screened and growth curves were determined. The transformation of Ri T-DNA was examined through PCR and baicalin content was examined through HPLC. RESULT: Hairy roots appeared in vitro 8 days after inoculation of S. viscidula with disarmed A. tumefaciens strain C58C1. After 24 days the frequency of stems explants was up to 81%. Transformation was confirmed by the amplification of rolC genes from the hairy roots of S. viscidula. The results demonstrated that rolC genes could be expressed in hairy roots of S. viscidula. Under the 36 days suspension culture of S. viscidula hairy roots in 1/2MS medium, dry weight of hairy root increased 17.42 times, the content and baicalin increased 21.60 and 25.56 times. CONCLUSION: The establishment of the culturing system of hairy root of S. viscidula provided a foundation for further industrial production of active drug component.

Comparisons of Scutellaria baicalensis, Scutellaria lateriflora and Scutellaria racemosa: genome size, antioxidant potential and phytochemistry.

The genus Scutellaria in the family Lamiaceae has over 350 species, many of which are medicinally active. One species, Scutellaria baicalensis, is one of the most widely prescribed plants in Traditional Chinese Medicine, used for neurological disorders, cancer and inflammatory diseases and has been the subject of detailed scientific study but little is known about the phytochemistry of other Scutellaria. The current study was designed to compare the medicinal phytochemistry of 3 species of Scutellaria used to treat neurological disorders. To accomplish this objective, the specific objectives were (a) to establish an in vitro collection of the South American native; S. racemosa, (b) to botanically characterize S. racemosa and (c) to compare the phytochemistry of S. racemosa with S. baicalensis and S. lateriflora. S. racemosa was established in vitro from wild populations in Florida. Botanically, S. racemosa is diploid with 18 chromosomes, and flow cytometry data indicated that S. baicalensis and S. racemosa have small nuclei with estimated small genomes (377 mbp and 411 mbp respectively). Antioxidant potential studies showed that there were no significant differences in the 3 Scutellaria species. Phytochemical analyses detected and quantified the flavonoids baicalin, baicalein, scutellarin, and wogonin as well as the human neurohormones melatonin and serotonin in leaf and stem tissues from S. baicalensis, S. lateriflora, and S. racemosa. These findings represent the first phytochemical analysis of S. racemosa and establish S. racemosa as a model system for study of medicinal plant secondary metabolism and as a potential source of important phytopharmaceuticals for treatment of human disease.