Antioxidant potential evaluation of polysaccharides from Camellia oleifera Abel in vitro and in vivo.

The extraction process, structural characterization and free radical scavenging ability of polysaccharides from Camellia oleifera have already been widely studied. However, the antioxidant activities are still lack of systematic experiments. In this study, we used Hep G2 cells and Caenorhabditis elegans to evaluate the antioxidant potential of polysaccharides that from C. oleifera flowers (P-CF), leaves (P-CL), seed cakes (P-CC) and fruit shells (P-CS). The results showed all these polysaccharides could protect cells from oxidative damage induced by t-BHP. The highest cell viabilities were 66.46 ± 1.36 % (P-CF), 55.2 ± 2.93 % (P-CL), 54.49 ± 1.29 % (P-CC) and 61.45 ± 1.67 % (P-CS), respectively. Studies have shown that four polysaccharides may protect cells from apoptosis by reducing ROS levels and maintaining MMP balance. Moreover, P-CF, P-CL, P-CC and P-CS increased the survival rate of C. elegans under thermal stress, which reduced the production of ROS by 56.1 ± 0.67 %, 59.37 ± 1.79 %, 16.63 ± 2.51 % and 27.55 ± 2.62 %, respectively. P-CF and P-CL showed stronger protective effects on C. elegans by increasing the nuclear entry rate of DAF-16 and stimulating the expression of SOD-3. Our study suggested that C. oleifera polysaccharides have the potential to develop into a natural supplement agent.

Structural features and antioxidant activities of polysaccharides from different parts of Codonopsis pilosula var. modesta (Nannf.) L. T. Shen.

In this study, three acidic polysaccharides from different plant parts of Codonopsis pilosula var. Modesta (Nannf.) L. T. Shen were obtained by ion exchange chromatography and gel filtration chromatography, and the yields of these three polysaccharides were different. According to the preliminary experimental results, the antioxidant activities of the polysaccharides from rhizomes and fibrous roots (CLFP-1) were poor, and was thus not studied further. Due to this the structural features of polysaccharides from roots (CLRP-1) and aerial parts (CLSP-1) were the object for this study and were structurally characterized, and their antioxidant activities were evaluated. As revealed by the results, the molecular weight of CLRP-1and CLSP-1 were 15.9 kDa and 26.4 kDa, respectively. The monosaccharide composition of CLRP-1 was Ara, Rha, Fuc, Xyl, Man, Gal, GlcA, GalA in a ratio of 3.8: 8.4: 1.0: 0.8: 2.4: 7.4: 7.5: 2.0: 66.7, and Ara, Rha, Gal, GalA in a ratio of 5.8: 8.9: 8.0: 77.0 in for CLSP-1. The results of structural elucidation indicated that both CLRP-1 and CLSP-1 were pectic polysaccharides, mainly composed of 1, 4-linked galacturonic acid with long homogalacturonan regions. Arabinogalactan type I and arabinogalactan type II were presented as side chains. The antioxidant assay in IPEC-J2 cells showed that both CLRP-1 and CLSP-1 promoted cell viability and antioxidant activity, which significantly increase the level of total antioxidant capacity and the activity of superoxide dismutase, catalase, and decrease the content of malondialdehyde. Moreover, CLRP-1 and CLSP-1 also showed powerful antioxidant abilities in Caenorhabditis elegans and might regulate the nuclear localization of DAF-16 transcription factor, induced antioxidant enzymes activities, and further reduced reactive oxygen species and malondialdehyde contents to increase the antioxidant ability of Caenorhabditis elegans. Thus, these finding suggest that CLRP-1 and CLSP-1 could be used as potential antioxidants.

Physicochemical Characterization of a Fern Polysaccharide from Alsophila spinulosa Leaf and Its Anti-Aging Activity in Caenorhabditis elegans.

Alsophila spinulosa, as a rare tree fern with potential medicinal value, has attracted extensive attention. Herein, the physicochemical properties, antioxidant and anti-aging activities of polysaccharide from A. spinulosa leaf (ALP) were investigated. ALP was composed of galactose, arabinose, glucose, rhamnose, galacturonic acid, mannose, and fucose. (1→), (1→6), and (1→2) bond types were the primary glycosidic bond in ALP. Surprisingly, ALP displayed the wonderful activity of antioxidant and anti-aging, including excellent scavenging ability against DPPH and ABTS radicals in vitro; prolonging the life span, improving activity of antioxidative enzymes (SOD and CAT), and decreasing the level of ROS, MDA in Caenorhabditis elegans. Meanwhile, ALP promoted DAF-16 to move into the nuclear. Overall, our results illustrated that ALP could be further developed as a functional food ingredient.

Melatonin Enhanced the Tolerance of Arabidopsis thaliana to High Light Through Improving Anti-oxidative System and Photosynthesis.

Land plants live in a crisis-filled environment and the fluctuation of sunlight intensity often causes damage to photosynthetic apparatus. Phyto-melatonin is an effective bioactive molecule that helps plants to resist various biotic and abiotic stresses. In order to explore the role of melatonin under high light stress, we investigated the effects of melatonin on anti-oxidative system and photosynthesis of Arabidopsis thaliana under high light. Results showed that exogenous melatonin increased photosynthetic rate and protected photosynthetic proteins under high light. This was mainly owing to the fact that exogenous melatonin effectively decreased the accumulation of reactive oxygen species and protected integrity of membrane and photosynthetic pigments, and reduced cell death. Taken together, our study promoted more comprehensive understanding in the protective effects of exogenous melatonin under high light.

Exogenous Melatonin Alleviates Oxidative Damages and Protects Photosystem II in Maize Seedlings Under Drought Stress.

The protective role of melatonin in plants against various abiotic stresses have been widely demonstrated, but poorly explored in organ-specific responses and the transmission of melatonin signals across organs. In this study, the effects of melatonin with the root-irrigation method and the leaf-spraying method on the antioxidant system and photosynthetic machinery in maize seedlings under drought stress were investigated. The results showed that drought stress led to the rise in reactive oxygen species (ROS), severe cell death, and degradation of D1 protein, which were mitigated by the melatonin application. The application of melatonin improved the photosynthetic activities and alleviated the oxidative damages of maize seedlings under the drought stress. Compared with the leaf-spraying method, the root-irrigation method was more effective on enhancing drought tolerance. Moreover, maize seedlings made organ-specific physiological responses to the drought stress, and the physiological effects of melatonin varied with the dosage, application methods and plant organs. The signals of exogenous melatonin received by roots could affect the stress responses of leaves, and the melatonin signals perceived by leaves also led to changes in physiological metabolisms in roots under the stress. Consequently, the whole seedlings coordinated the different parts and made a systemic acclimation against the drought stress. Melatonin as a protective agent against abiotic stresses has a potential application prospect in the agricultural industry.

Exogenous melatonin enhances salt stress tolerance in maize seedlings by improving antioxidant and photosynthetic capacity.

Melatonin (N-acetyl-5-methoxytryptamine) is an important biological hormone in many abiotic stress responses and developmental processes. In this study, the protective roles of melatonin were investigated by measuring the antioxidant defense system and photosynthetic characteristics in maize under salt stress. The results indicated that NaCl treatment led to the decrease in plant growth, chlorophyll contents and photochemical activity of photosystem II (PSII). However, the levels of reactive oxygen species increased significantly under salt stress. Meanwhile, we found that application of exogenous melatonin alleviated reactive oxygen species burst and protected the photosynthetic activity in maize seedlings under salt stress through the activation of antioxidant enzymes. In addition, 100 µM melatonin-treated plants showed high photosynthetic efficiency and salinity. Immunoblotting analysis of PSII proteins showed that melatonin application alleviated the decline of 34 kDa PSII reaction center protein (D1) and the increase of PSII subunit S protein. Taken together, our study promotes more comprehensive understanding in the protective effects of exogenous melatonin in maize under salt stress, and it may be involved in activation of antioxidant enzymes and regulation of PSII proteins.

Effects of Melatonin on Anti-oxidative Systems and Photosystem II in Cold-Stressed Rice Seedlings.

Melatonin (N-acetyl-5-methoxytryptamine) plays important role in multiple plant developmental processes and stress responses. We investigated the possible mediatory role of melatonin in growth, photosynthesis, and the response to cold stress in rice by using three different experiments: soaking seed; immersing roots, and spraying to leaves with 0, 20, or 100 µM melatonin. After 6 days of cold stress, the growth of rice seedlings was significantly inhibited, but this inhibition was alleviated by exogenous melatonin. Furthermore, exogenous melatonin pretreatment alleviated the accumulation of reactive oxygen species, malondialdehyde and cell death induced by cold stress. Melatonin pretreatment also relieved the stress-induced inhibitions to photosynthesis and photosystem II activities. Further investigations showed that, antioxidant enzyme activities and non-enzymatic antioxidant levels were increased by melatonin pretreatments. The treatment methods of seed soaking and root immersion were more effective in improving cold stress resistance than the spraying method. The results also indicated the dose-dependent response of melatonin on rice physiological, biochemical, and photosynthetic parameters.

Phylogeny and differentiation of the St genome in Elymus L. sensu lato (Triticeae; Poaceae) based on one nuclear DNA and two chloroplast genes.

BACKGROUND: Hybridization and polyploidization can be major mechanisms for plant evolution and speciation. Thus, the process of polyploidization and evolutionary history of polyploids is of widespread interest. The species in Elymus L. sensu lato are allopolyploids that share a common St genome from Pseudoroegneria in different combinations with H, Y, P, and W genomes. But how the St genome evolved in the Elymus s. l. during the hybridization and polyploidization events remains unclear. We used nuclear and chloroplast DNA-based phylogenetic analyses to shed some light on this process. RESULTS: The Maximum likelihood (ML) tree based on nuclear ribosomal internal transcribed spacer region (nrITS) data showed that the Pseudoroegneria, Hordeum and Agropyron species served as the St, H and P genome diploid ancestors, respectively, for the Elymus s. l. polyploids. The ML tree for the chloroplast genes (matK and the intergenic region of trnH-psbA) suggests that the Pseudoroegneria served as the maternal donor of the St genome for Elymus s. l. Furthermore, it suggested that Pseudoroegneria species from Central Asia and Europe were more ancient than those from North America. The molecular evolution in the St genome appeared to be non-random following the polyploidy event with a departure from the equilibrium neutral model due to a genetic bottleneck caused by recent polyploidization. CONCLUSION: Our results suggest the ancient common maternal ancestral genome in Elymus s. l. is the St genome from Pseudoroegneria. The evolutionary differentiation of the St genome in Elymus s. l. after rise of this group may have multiple causes, including hybridization and polyploidization. They also suggest that E. tangutorum should be treated as C. dahurica var. tangutorum, and E. breviaristatus should be transferred into Campeiostachys. We hypothesized that the Elymus s. l. species origined in Central Asia and Europe, then spread to North America. Further study of intraspecific variation may help us evaluate our phylogenetic results in greater detail and with more certainty.

Characterization, antioxidant and antitumor activities of polysaccharides from Salvia miltiorrhiza Bunge.

Polysaccharides were extracted from Salvia miltiorrhiza Bunge using response surface methodology (RSM) with ultrasonication. A Box-Behnken design was used to optimize the extraction parameters to maximize the polysaccharide extraction yield. The polysaccharide SMP-U1 was isolated and characterized; then the antioxidant and antiproliferation activities were evaluated in vitro. The modified optimal conditions were an ultrasonic power of 180 W, an extraction temperature of 54°C, and an extraction time of 32 min, achieving an extraction yield of 40.54±0.25%. The results indicate that SMP-U1 has significant antioxidant activity, scavenging the free radical 2,2-diphenyl-1-picrylhydrazyl. It has also exhibited effect on the proliferation of human breast carcinoma cells Bcap-37 and human esophageal carcinoma cells Eca-109, especially at a concentration of 0.30 mg/mL. In conclusion, SMP-U1 has remarkable in vitro antioxidant and antiproliferation activity, and has potential for application as a natural antioxidant or antitumor agent.

Evolutionary pattern of rDNA following polyploidy in Leymus (Triticeae: Poaceae).

Ribosomal ITS polymorphism and its ancestral genome origin of polyploid Leymus were examined to infer the evolutionary outcome of rDNA gene following allopolyploid speciation and to elucidate the geographic pattern of ITS variation. The results demonstrated that different polyploids have experienced varying fates, including maintenance or homogenization of divergent arrays, occurrence of chimeric repeats and potential pseudogenes. Our data suggested that (1) the Ns, P/F, and St genomic types in Leymus were originated from Psathyrostachys, Agropyron/Eremopyrum, and Pseudoroegneria, respectively; (2) the occurrence of a higher proportion of Leymus species with predominant uniparental rDNA type might associate with the segmental allopolyploid origin, nucleolar dominance of alloploids, and rapid radiation of Leymus; (3) maintenance of multiple parental ITS types in allopolyploid might result from long generation times associated to vegetative multiplication, number and chromosomal location of ribosomal loci and/or recurrent hybridization; (4) the rDNA genealogical structure of Leymus species might associate with the geographic origins; and (5) ITS sequence clade shared by Leymus species from Central Asia, North America, and Nordic might be an outcome of ancestral ITS homogenization. Our results shed new light on understanding evolutionary outcomes of rDNA following allopolyploid speciation and geographic isolation.

[Molecular cloning of squalene synthase gene form Paris polyphylla and its expression in Escherichia coli].

OBJECTIVE: To clone the cDNA sequence of squalene synthase gene from Paris polyphylla, and characterize the biological features of the obtained SQS. METHOD: Using homology cloning and RACE technique, a full-length cDNA sequence of PpSQS gene was isolated from P. polyphylla. The obtained sequence was analyzed by bioinformatics softwares. A plasmid [named pET-30b (+)-PpSQS] was constructed for prokaryotic expression the recombinant PpSQS. RESULT: The full-length cDNA of PpSQS gene is 1 498 bp, which contains a 1 212 bp ORF. Sequence analysis indicated that PpSQS encoded 403 amino acids residues with a calculated molecular weight (MW) of 46.36 kDa and an isoelectric point (pI) of 6.83. SDS-PAGE results showed that the recombinant PpSQS was expressed in Escherichia coli BL21 (DE3) by inducing with 1 mmol x L(-1) IPTG. CONCLUSION: The full-length cDNA sequence of PpSQS gene was obtained from P. polyphylla, and its molecular features were consisted with classic SQS in plant. The recombinant PpSQS was successfully expressed in E. coli.

Evolutionary history of Triticum petropavlovskyi Udacz. et Migusch. inferred from the sequences of the 3-phosphoglycerate kinase gene.

Single- and low-copy genes are less likely to be subject to concerted evolution. Thus, they are appropriate tools to study the origin and evolution of polyploidy plant taxa. The plastid 3-phosphoglycerate kinase gene (Pgk-1) sequences from 44 accessions of Triticum and Aegilops, representing diploid, tetraploid, and hexaploid wheats, were used to estimate the origin of Triticum petropavlovskyi. Our phylogenetic analysis was carried out on exon+intron, exon and intron sequences, using maximum likelihood, Bayesian inference and haplotype networking. We found the D genome sequences of Pgk-1 genes from T. petropavlovskyi are similar to the D genome orthologs in T. aestivum, while their relationship with Ae. tauschii is more distant. The A genome sequences of T. petropavlovskyi group with those of T. polonicum, but its Pgk-1 B genome sequences to some extent diverge from those of other species of Triticum. Our data do not support for the origin of T. petropavlovskyi either as an independent allopolyploidization event between Ae. tauschii and T. polonicum, or as a monomendelian mutation in T. aestivum. We suggest that T. petropavlovskyi originated via spontaneous introgression from T. polonicum into T. aestivum. The dating of divergence among T. polonicum, T. petropavlovskyi, T. carthlicum, T. turgidum, and T. compactum indicates an age of 0.78 million years [corrected].

Phylogenetic relationships and Y genome origin in Elymus L. sensu lato (Triticeae; Poaceae) based on single-copy nuclear Acc1 and Pgk1 gene sequences.

To estimate the origin and genomic relationships of the polyploid species within Elymus L. sensu lato, two unlinked single-copy nuclear gene (Acc1 and Pgk1) sequences of eighteen tetraploids (StH and StY genomes) and fourteen hexaploids (StStH, StYP, StYH, and StYW genomes) were analyzed with those of 35 diploid taxa representing 18 basic genomes in Triticeae. Sequence and phylogenetic analysis suggested that: (1) the St, H, W, and P genomes were donated by Pseudoroegneria, Hordeum, Australopyrum, and Agropyron, respectively, while the Y genome is closely related to the Xp genome in Peridictyon sanctum; (2) different hexaploid Elymus s.l. species may derived their StY genome from different StY genome tetraploid species via independent origins; (3) due to incomplete lineage sorting and/or hybridization events, the genealogical conflict between the two gene trees suggest introgression involving some Elymus s.l. species, Pseudoroegneria, Agropyron and Aegilops/Triticum; (4) it is reasonable to recognize the StH genome species as Elymus sensu stricto, the StY genome species as Roegneria, the StYW genome species as Anthosachne, the StYH genome species as Campeiostachys, and the StYP genome species as Kengyilia. The occurrence of multiple origin and introgression could account for the rich diversity and ecological adaptation of Elymus s.l. species.

Potential use of DNA barcoding for the identification of Salvia based on cpDNA and nrDNA sequences.

An effective DNA marker for authenticating the genus Salvia was screened using seven DNA regions (rbcL, matK, trnL-F, and psbA-trnH from the chloroplast genome, and ITS, ITS1, and ITS2 from the nuclear genome) and three combinations (rbcL+matK, psbA-trnH+ITS1, and trnL-F+ITS1). The present study collected 232 sequences from 27 Salvia species through DNA sequencing and 77 sequences within the same taxa from the GenBank. The discriminatory capabilities of these regions were evaluated in terms of PCR amplification success, intraspecific and interspecific divergence, DNA barcoding gaps, and identification efficiency via a tree-based method. ITS1 was superior to the other marker for discriminating between species, with an accuracy of 81.48%. The three combinations did not increase species discrimination. Finally, we found that ITS1 is a powerful barcode for identifying Salvia species, especially Salvia miltiorrhiza.

Evolutionary dynamics of the Pgk1 gene in the polyploid genus Kengyilia (Triticeae: Poaceae) and its diploid relatives.

The level and pattern of nucleotide variation in duplicate gene provide important information on the evolutionary history of polyploids and divergent process between homoeologous loci within lineages. Kengyilia is a group of allohexaploid species with the StYP genomic constitutions in the wheat tribe. To investigate the evolutionary dynamics of the Pgk1 gene in Kengyilia and its diploid relatives, three copies of Pgk1 homoeologues were isolated from all sampled hexaploid Kengyilia species and analyzed with the Pgk1 sequences from 47 diploid taxa representing 18 basic genomes in Triticeae. Sequence diversity patterns and genealogical analysis suggested that (1) Kengyilia species from the Central Asia and the Qinghai-Tibetan plateau have independent origins with geographically differentiated P genome donors and diverged levels of nucleotide diversity at Pgk1 locus; (2) a relatively long-time sweep event has allowed the Pgk1 gene within Agropyron to adapt to cold climate triggered by the recent uplifts of the Qinghai-Tibetan Plateau; (3) sweep event and population expansion might result in the difference in the d(N)/d(S) value of the Pgk1 gene in allopatric Agropyron populations, and this difference may be genetically transmitted to Kengyilia lineages via independent polyploidization events; (4) an 83 bp MITE element insertion has shaped the Pgk1 loci in the P genome lineage with different geographical regions; (5) the St and P genomes in Kengyilia were donated by Pseudoroegneria and Agropyron, respectively, and the Y genome is closely related to the Xp genome of Peridictyon sanctum. The interplay of evolutionary forces involving diverged natural selection, population expansion, and transposable events in geographically differentiated P genome donors could attribute to geographical differentiation of Kengyilia species via independent origins.

Genetic diversity and phylogeny in Hystrix (Poaceae, Triticeae) and related genera inferred from Giemsa C-banded karyotypes.

The phylogenetic relationships of 15 taxa from Hystrix and the related genera Leymus (NsXm), Elymus (StH), Pseudoroegneria (St), Hordeum (H), Psathyrostachys (Ns), and Thinopyrum (E) were examined by using the Giemsa C-banded karyotype. The Hy. patula C-banding pattern was similar to those of Elymus species, whereas C-banding patterns of the other Hystrix species were similar to those of Leymus species. The results suggest high genetic diversity within Hystrix, and support treating Hy. patula as E. hystrix L., and transferring Hy. coreana, Hy. duthiei ssp. duthiei and Hy. duthiei ssp. longearistata to the genus Leymus. On comparing C-banding patterns of Elymus species with their diploid ancestors (Pseudoroegneria and Hordeum), there are indications that certain chromosomal re-arrangements had previously occurred in the St and H genomes. Furthermore, a comparison of the C-banding patterns of the Hystrix and Leymus species with the potential diploid progenitors (Psathyrostachys and Thinopyrum) suggests that Hy. coreana and some Leymus species are closely related to the Ns genome of Psathyrostachys, whereas Hy. duthiei ssp. duthiei, Hy. duthiei ssp. longearistata and some of the Leymus species have a close relationship with the E genome. The results suggest a multiple origin of the polyploid genera Hystrix and Leymus.

Genetic diversity and phylogeny in Hystrix (Poaceae, Triticeae) and related genera inferred from Giemsa C-banded karyotypes

The phylogenetic relationships of 15 taxa from Hystrix and the related genera Leymus (NsXm), Elymus (StH), Pseudoroegneria (St), Hordeum (H), Psathyrostachys (Ns), and Thinopyrum (E) were examined by using the Giemsa C-banded karyotype. The Hy. patula C-banding pattern was similar to those of Elymus species, whereas C-banding patterns of the other Hystrix species were similar to those of Leymus species. The results suggest high genetic diversity within Hystrix, and support treating Hy. patula as E. hystrix L., and transferring Hy. coreana, Hy. duthiei ssp. duthiei and Hy. duthiei ssp. longearistata to the genus Leymus. On comparing C-banding patterns of Elymus species with their diploid ancestors (Pseudoroegneria and Hordeum), there are indications that certain chromosomal re-arrangements had previously occurred in the St and H genomes. Furthermore, a comparison of the C-banding patterns of the Hystrix and Leymus species with the potential diploid progenitors (Psathyrostachys and Thinopyrum) suggests that Hy. coreana and some Leymus species are closely related to the Ns genome of Psathyrostachys, whereas Hy. duthiei ssp. duthiei, Hy. duthiei ssp. longearistata and some of the Leymus species have a close relationship with the E genome. The results suggest a multiple origin of the polyploid genera Hystrix and Leymus.

[Principal component and cluster analysis of trace elements in Chinese herb Salvia miltiorrhiza and its relative species].

The contents of trace elements, including copper, zinc, iron, magnesium, calcium, chromium, lead, molybdenum, manganese, and cadmium in Chinese traditional herb S. miltiorrhiza and its relative species such as S. miltiorrhiza f. alba, S. evansiana, S. yunnanensis, S. przewarlskii, S. cavaleriei, S. cavaleriei var. simplicifolia, S. roborowskii, S. omeiana, S. tricuspis, S. brevilabra and S. cynica were determined by atomic absorption spectroscopy. The principal components analysis of SPSS was applied to the study of characteristic elements in S. miltiorrhiza and its relative species. Three principal components which accounted for 79.3% of total variance were extracted from the original data. The first factor accounted for 49.6% of the total variance, which means that iron, manganese, copper, zinc, cadmium and lead are the characteristic elements in S. miltiorrhiza and its relative species. The results of cluster analysis show that the samples could cluster reasonably into two groups. The samples of S. miltiorrhiza from different regions were classified into one group except S. miltiorrhiza from Zhejiang and S. miltiorrhiza f. alba. The other Salvia species were classified into another group except S. cavaleriei. The S. miltiorrhiza and other Salvia species can be distinguished by this method, whose accuracy of classification is 90%. The cluster analysis based on the contents of trace element in S. miltiorrhiza and its relative species provided a quick, accurate and simple method for authentication of herb Salvia miltiorrhiza.

[PCR-RFLP analysis on Roegneria, Elymus, Hystrix and Kengyilia in Triticeae (Poaceae)].

PCR-RFLP of Plasmon on 23 species of Roegneria, Elymus, Hystrix and Kengyilia and 1 outgroup (Triticum aestivum L.) was carried out. The segments amplified with 3 cpDNA and 3 mtDNA universal primers were digested by 15 restriction endonucleases. Among 329 bands produced, 304 were polymorphic (92.4%). PCR-RFLP results showed that polymorphisms existed between the four genera and between the species in Roegneria, Elymus, Hystrix and Kengyilia, and genetic similarity coefficient was high. Roegneria grandis, R. aristiglumis, R. elytrigioides, R. alashanica and R. magnicaespes were grouped, while R. caucasica, R. ciliaris, R. amurensis and R. japonensis clustered together. Species of Kengyilia clustered independently. Five species of Elymus, 3 of Hystrix and Roegneria kamoji grouped together. The results suggested Kengyilia was a valid genus. As more cytological and molecular data are available, the taxonomic status of Roegneria, Elymus and Hystrix will be recognized.

[Studies on chromosome numbers of Salvia miltiorrhiza, S. flava and S. evansiana].

OBJECTIVE: To study the numbers of chromosome in Chinese herb Salvia miltiorrhiza from 7 provinces in China, and S. flava as well as S. evansiana from Yunnan province in China. METHOD: The young root was treated with the mixture of ice and water for 24 h, fixed with Carony's fixative for 6-12 h. After differentiating for 10-12 min with 1 mol x L(-1) hydrochloric acid at 60 'C and staining with carbol fuchsin,the section was observed under microscope. RESULT: Chromosome numbers of S. miltiorrhiza and S.flava were 2n = 2x = 16. The numbers of S. evansiana were 2n = 4x = 32. The basic numbers of the chromosomes were x = 8. And tetraploids were observed in S. miltiorrhiza from Sichuan provices and Shandong provices. CONCLUSION: The basic number of the chromosomes are x = 8. The chromosome numbers of S. miltiorrhiza, S.flava and S. evansiana are 16,16 and 32 respectively. As the chromosomes are the small or micro-small ones, it is difficult to use them for karyotype.