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1.
Electron J Biotechnol ; 49: 50-55, Jan. 2021. tab, graf
Article in English | LILACS | ID: biblio-1291649

ABSTRACT

BACKGROUND: Euphorbia fischeriana Steud is a very important medicinal herb and has significant medical value for healing cancer, edema and tuberculosis in China. The lack of molecular markers for Euphorbia fischeriana Steud is a dominant barrier to genetic research. For the purpose of developing many simple sequence repeat (SSR) molecular markers, we completed transcriptome analysis with the Illumina HiSeq 2000 platform. RESULTS: Approximately 9.1 million clean reads were acquired and then assembled into approximately 186.3 thousand nonredundant unigenes, 53,146 of which were SSR-containing unigenes. A total of 76,193 SSR loci were identified. Of these SSR loci, 28,491 were detected at the terminal position of ESTs, which made it difficult to design SSR primers for these SSR-containing sequences, and the residual SSRs were thus used to design primer pairs. Analyzing the results of these markers revealed that the mononucleotide motif A/T (44,067, 57.83% of all SSRs) was the most abundant, followed by the dinucleotide type AG/CT (9430, 12.38%). Using 100 randomly selected primer pairs, 77 primers were successfully amplified in Euphorbia fischeriana Steud, and 79 were successfully amplified in three other related species. The markers developed displayed relatively high quality and cross-species transferability. CONCLUSIONS: The large number of EST-SSRs exploited successfully in Euphorbia fischeriana Steud for the first time could provide genetic information for research on linkage maps, variety identification, genetic diversity analysis, and molecular marker-assisted breeding.


Subject(s)
Euphorbia/genetics , High-Throughput Nucleotide Sequencing/methods , Plants, Medicinal , Genetic Variation , Genetic Markers
2.
Article in Chinese | WPRIM | ID: wpr-908353

ABSTRACT

High-throughput sequencing technology has been widely used to diagnose neonatal genetic diseases.Compared with conventional next-generation sequencing technology, the sequencing process of rapid high-throughput sequencing is more complete and with shorter sequencing time.The rapid high-throughput sequencing can be used for timely adjustment of clinical management.This review systematically analyzes the application of rapid high-throughput sequencing technology in the diagnosis of neonatal genetic diseases, and provides a reference for its routine clinical application.

3.
Article in Chinese | WPRIM | ID: wpr-906439

ABSTRACT

Objective:To explore the differences in rhizosphere microbial community structure between <italic>Fusarium</italic> wilt-infected and healthy <italic>Chrysanthemum morifolium </italic>plants<italic>.</italic> Method:The rhizosphere soils of diseased and healthy<italic> C. morifolium </italic>plants were sampled and subjected to high-throughput 16S ribosomal DNA (rDNA) and internal transcribed spacer (ITS) sequencing, to identify the microbial community structure including bacteria and fungi. Result:<italic>Fusarium</italic> wilt reduced the bacterial abundance and diversity but had no significant effect on fungal alpha-diversity.The proportions of Acidobacteria, Gemmatimonadetes, and Nitrospirae in rhizosphere soil of healthy <italic>C.morifolium</italic> plants were higher than those of diseased plants, while the proportions of Proteobacteria and Bacteroidetes were lower(<italic>P</italic><0.05). <italic>Fusarium</italic> fungi accounted for 27.49%, 14.53%, and 11.94% in diseased plants whereas 0.47%, 1.01%, and 0.67% in healthy plants.Pathogenic bacteria <italic>Pectobacterium</italic> and <italic>Dickeya</italic> were enriched in rhizosphere soil of diseased plants. The abundances of nitrifying, detoxifying, and photosynthetic bacteria in rhizosphere soil of healthy plants were higher than those of diseased plants. Conclusion:<italic>Fusarium</italic> wilt reduces the bacterial richness and diversity and triggers the enrichment of massive <italic>Fusarium</italic> fungi, <italic>Pectobacterium</italic>, and <italic>Dickeya</italic>. The proportion of beneficial bacteria in rhizosphere soil of healthy plants is significantly higher than that of diseased plants.

4.
Article in Chinese | WPRIM | ID: wpr-906346

ABSTRACT

Objective:To explore the potential molecular mechanism of corylin in the treatment of lung cancer. Method:A549 cells were treated with different concentrations of corylin, and their proliferation was detected using methye thiazolye telrazlium (MTT) reagent. Then the trend analysis and gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were conducted to screen the key genes and pathways of corylin against A549 cell proliferation, followed by the verification of sequencing results by real-time polymerase chain reaction (Real-time PCR). Result:Corylin inhibited the proliferation of A549 cells and regulated the expression of 4 364 genes in cells. The trend analysis revealed that these genes were clustered into 20 distinct modules, among which four were significantly down-regulated, suggesting that corylin exerted the anti-proliferation effect by inhibiting the expression of some genes. The inter-group comparison of differentially expressed genes (DEGs) showed that the elevation in the concentration of corylin resulted in more down-regulated genes but weakened proliferation, consistent with the findings by trend analysis. The GO and KEGG enrichment analysis of 278 DEGs in the high-dose corylin group demonstrated that corylin mainly changed the cellular and metabolic processes, which was attributed to its regulation of steroid biosynthesis, fatty acid metabolism, biosynthesis of unsaturated fatty acids, synthesis and degradation of ketone bodies, and steroid hormone biosynthesis. The Real-time PCR results confirmed that corylin down-regulated the mRNA expression levels of LSS, stearoyl-CoA desaturase (SCD), 3-hydroxy-3-methylglutaryl-CoA synthase 1 (HMGCS1), but up-regulated the mRNA expression of recombinant human angiopoietin-like protein 4 (ANGPTL4), basically consistent with the transcriptomics results. Conclusion:Corylin inhibits A549 cell proliferation and alleviates lung cancer by targeting the related genes in lipid metabolism pathways.

5.
Article in Chinese | WPRIM | ID: wpr-906080

ABSTRACT

Objective:To compare the effects of Baiyaojian before and after fermentation on intestinal flora and expression of Occludin and zonula occludens protein-1 (ZO-1) in intestinal mucosa of mice with ulcerative colitis (UC), and to explore the mechanism of Baiyaojian and Galla Chinensis in the treatment of UC. Method:Totally 50 mice were randomly divided into 5 groups with 10 mice in each group, one group was randomly selected as blank group, and the other 4 groups were treated with dextran sodium sulfate (DSS) to induce UC model. After modeling, mice in the blank group and model group were given normal saline, and treatment groups were given Mesalazine (0.8 g·kg<sup>-1</sup>), Galla Chinensis decoction (1.8 g·kg<sup>-1</sup>) and Baiyaojian decoction (2.7 g·kg<sup>-1</sup>) by intragastric administration for 7 days. The 16S rRNA sequencing technology was used to detect the changes of intestinal flora in mouse feces. The histopathological changes of colon tissue were observed by hematoxylin-eosin (HE) staining, and the expression of Occludin and ZO-1 in colon tissue of mice were compared by immunohistochemistry. Result:Compared with the blank group, the abundance and diversity of intestinal flora in UC mice were significantly decreased, and the colonic tissue was thickened with congestion and obvious ulcers, and the expression levels of Occludin and ZO-1 were significantly decreased (<italic>P</italic><0.01). After treatment with Galla Chinensis and Baiyaojian, the abundance and diversity of flora were improved. At the phylum level, relative abundance of Firmicutes, Proteobacteria and Actinobacteria increased significantly (<italic>P</italic><0.01), while the relative abundance of Bacteroidetes decreased significantly (<italic>P</italic><0.01) in Galla Chinensis group. In Baiyaojian group, the relative abundance of Proteobacteria and Actinobacteria increased significantly (<italic>P</italic><0.01), while the relative abundance of Firmicutes increased and the relative abundance of Bacteroidetes decreased, but there was no significant difference. At the genus level, the relative abundance of <italic>Bacteroides</italic>, <italic>Allobaculum </italic>and <italic>Ruminococcus</italic> decreased significantly (<italic>P</italic><0.01), the relative abundance of <italic>Roseburia</italic>, <italic>Prevotella</italic>, <italic>Oscillospira</italic> and <italic>Paraprevotella</italic> increased significantly (<italic>P</italic><0.05, <italic>P</italic><0.01) in Galla Chinensis group. In Baiyaojian group, the relative abundance of <italic>Bacteroides</italic> and <italic>Allobaculum</italic> decreased significantly (<italic>P</italic><0.05, <italic>P</italic><0.01), and the relative abundance of <italic>Prevotella</italic>, <italic>Oscillospira</italic>, <italic>Roseburia</italic> and <italic>Ruminococcus</italic> increased significantly (<italic>P</italic><0.01). Compared with model group, colon tissue of Galla Chinensis group and Baiyaojian group was recovered obviously, congestion was alleviated, only scattered ulcers were seen. The expression of Occludin and ZO-1 increased, and the expression level of Baiyaojian group was higher than that of Galla Chinensis group. Conclusion:The effect of Baiyaojian is better than Galla Chinensis in the treatment of UC. The mechanism may be through regulating the abundance and diversity of intestinal flora, improving the disorder of intestinal flora and increasing the expression of ZO-1 and Occludin and protecting the intestinal mucosal barrier function for alleviating intestinal inflammation.

6.
China Occupational Medicine ; (6): 121-126, 2021.
Article in Chinese | WPRIM | ID: wpr-923223

ABSTRACT

OBJECTIVE: To explore the mechanism of bone marrow mesenchymal stem cells(BMSCs) in alleviating silica-induced lung injury in mice. METHODS: Ten specific pathogen free healthy male C57 BL/6 mice were selected for isolating BMSCs and bone marrow macrophages(BMDMs). Transwell chamber was used, BMDMs were inoculated onto the upper chamber and BMSCs in the lower chamber. We divided them into sequencing control group and silica(SiO_2) exposure group. All cells were pre-stimulated with 50 μg/L mass concentration lipopolysaccharide for 4 hours. In the SiO_(2 ) group, 250 mg/L mass concentration SiO_2 was added to the upper chamber of transwell and cultured for 16 hours. Total RNA was extracted from the BMSCs collected from the lower chamber. HiSeq/MiSeq high-throughput sequencing technology was used to detect the BMSCs RNA paired-end sequencing. Transcriptome sequencing data was obtained and bioinformatics analysis was performed. Another 12 specific pathogen free healthy male C57 BL/6 mice were randomly divided into control group and experimental group. All mice received one intra-tracheal injection of 20.0 μL(250 g/L mass concentration) of silica dust suspension. After 6 hours, the mice in the control group was given 500.0 μL of 0.9% sodium chloride solution and mice in the experimental group was given 500.0 μL of BMSCs suspension(cell density 1×10~9/L) by tail vein infusion.Mice were sacrificed 12 hours later. The relative mRNA expression of interleukin(IL)-1 Ra, IL-10, tumor necrosis factor stimulating gene 6(TSG-6) and prostaglandin E2(PGE2) in lung tissues of mice were measured by quantitative real-time PCR(q-PCR). Meanwhile, BMDMs and BMSCs transwell co-culture models were established. The cells were divided into 5 groups: BMSCs group, BMSCs+BMDMs group, BMSCs+BMDMs+ lipopolysaccharide(LPS) group, 50 mg/L SiO_2 group, and 100 mg/L SiO_2 group. After 16 hours of corresponding SiO_2 stimulation, BMSCs of each group were collected and the relative mRNA expression levels of IL-1 Ra, IL-10, TSG-6, and PGE2 in the cells were detected by q-PCR. RESULTS: Compared with sequencing control group, BMSCs co-cultured with SiO_2 had 19 genes up-regulated and 21 genes down-regulated, including 10 genes up-regulated for more than 2.0-fold. The relative mRNA expression of IL-1 Ra, IL-10, PGE2 and TSG-6 in the lung tissue of mice increased in the experimental group than that of the control group(all P<0.05). The relative mRNA expression of TSG-6 increased by 37.5 times higher than that of the control group. Compared with the BMSCs+BMDM+LPS group, the level of TSG-6 mRNA relative expression increased in both the 50 mg/L SiO_2 group and the 100 mg/L SiO_2 group(all P<0.05). CONCLUSION: TSG-6 could be the key factor of BMSCs that can attenuate silica-induced lung injury.

7.
Article in English | WPRIM | ID: wpr-922404

ABSTRACT

OBJECTIVES@#To investigate the diversity of peripheral blood T cell receptor (TCR) β chain complementarity-determining region 3 (CDR3) based on immune repertoire sequencing in neonates with sepsis and the possible pathogenesis of neonatal sepsis.@*METHODS@#A total of 12 neonates with sepsis were enrolled as the case group, and 9 healthy full-term infants, matched for gestational age, birth weight, and age, were enrolled as the control group. Omega nucleic acid purification kit (SQ blood DNA Kit II) was used to extract DNA from peripheral blood samples, TCR β chain CDR3 was amplified by multiplex PCR, and then high-throughput sequencing was performed for the products to analyze the diversity of TCR β chain CDR3 and the difference in expression.@*RESULTS@#The length and type of TCR β chain CDR3 were similar between the case and control groups, and Gaussian distribution was observed in both groups. With D50 and Shannon-Wiener index as the evaluation indices for diversity, the case group had a significantly lower diversity of TCR β chain CDR3 than the control group (@*CONCLUSIONS@#There is a significant change in the diversity of TCR β chain CDR3 in the peripheral blood of neonates with sepsis, suggesting that it might be associated with the immune pathogenesis of neonatal sepsis.


Subject(s)
Complementarity Determining Regions/genetics , High-Throughput Nucleotide Sequencing , Humans , Multiplex Polymerase Chain Reaction , Neonatal Sepsis , Receptors, Antigen, T-Cell, alpha-beta/genetics
8.
Article in Chinese | WPRIM | ID: wpr-912471

ABSTRACT

Inherited bleeding and thrombotic disorders (BTD) are a group of heterogeneous diseases related to the coagulation system, platelet function and fibrinolytic system. In addition to the common BTD, it is difficult to diagnose such patients by routine laboratory tests, and special laboratory tests are often required to confirm the diagnosis. Therefore, the diagnosis and treatment of patients may be delayed, or even life-threatening. With the increasing use of high-throughput sequencing (HTS) technology in clinical practice, the diagnosis and differential diagnosis of BTD have made great progress.

9.
Chinese Journal of Nephrology ; (12): 809-816, 2021.
Article in Chinese | WPRIM | ID: wpr-911903

ABSTRACT

Objective:To explore the mechanism of cisplatin-induced renal interstitial fibrosis and provide a new idea for the prevention and treatment of renal interstitial fibrosis.Methods:Eight-week-old male C57BL/6 mice (specific pathogen-free) were used to carry out the experiment. The mice were divided into cisplatin group (10 mg/kg, n=6) and saline group ( n=6) with intraperitoneal injection on day 0, 7 and 21, and sacrificed on day 28. The kidney tissues were collected for RNA Illumina high-throughput sequencing, real-time PCR, Western blotting, Masson staining and bioinformatics analysis. Results:Through real-time PCR, Western blotting and Masson staining, a mouse model with cisplatin-induced renal interstitial fibrosis was successfully established. Through RNA Illumina high-throughput sequencing, 387 long noncoding RNA (lncRNA) and 2 427 mRNA were differently expressed between cisplatin group and saline group. The expression of the top two lncRNA was confirmed by real-time PCR with the same tendency as RNA sequencing. Complement C3 was found to be at the top among the different expressed mRNA by RNA sequencing. Several terms related to immunity were found to be within the top 20 terms through Gene Ontology (GO) enrichment analysis. Systemic lupus erythematous pathway (ko05322, Q=3.4E-17), including the complement cascade pathway, was found to be the top pathway through Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. The mRNA expression levels of C1q, C2, C3 and C4 were up-regulated remarkably in the cisplatin group by RNA sequencing than those in saline group (all P<0.05) and confirmed by real-time PCR. Conclusions:Renal interstitial fibrosis can be induced by intraperitoneal injection of cisplatin periodically in mice, with complement cascade pathway activation in the diseased kidney.

10.
Article in Chinese | WPRIM | ID: wpr-888068

ABSTRACT

In this study, the colonization, diversity and relative abundance of arbuscular mycorrhizal fungi(AMF) in the roots of Panax quinquefolius in different habitats of Shandong province were analyzed by staining-microscopy and high-throughput sequencing. The data were analyzed by bioinformatics tools and statistical software. The results showed that the roots of P. quinquefolius in different habitats were colonized by AMF with different rates and intensities. The AMF in roots of P. quinquefolius belong to three genera, three families, three orders, one class and one phylum. At the level of order, the AMF mainly included Paraglomerales(52.48%), Glomerales(25.60%) and Archaeosporales(3.08%). At the level of family, the AMF were dominated by Paraglomeraceae(52.48%), Glomeraceae(18.94%) and Claroideoglomeraceae(3.05%). At the level of genus, Paraglomus(51.46%), Glomus(20.01%) and Claroideoglomus(3.52%) accounted for a large proportion, of which Paraglomus and Glomus were dominant. Cluster analysis showed that the AMF in roots of P. quinquefolius with close geographical locations could be clustered together. In this study, the diversity and dominant germplasm resources of AMF in roots of P. quinquefolius cultivated in the main producing areas were identified, which provi-ded basic data for revealing the quality formation mechanism of P. quinquefolius medicinal materials from the perspective of environment.


Subject(s)
Fungi , Glomeromycota , Humans , Mycorrhizae/genetics , Panax , Plant Roots , Soil Microbiology
11.
Article in Chinese | WPRIM | ID: wpr-888050

ABSTRACT

To obtain the difference of the fungal and bacterial community diversity between wild Cordyceps sinensis, artificial C. sinensis and their habitat soil, Illmina Hiseq high-throughput sequencing technology was applied. The results show that Proteobacteria was the dominant bacterial phylum in C. sinensis, Actinobacteria was the dominant bacterial phylum in soil microhabitat, Ophiocordyceps sinensis was the predominant dominant fungus of C. sinensis. The α diversity analysis showed that the fungal diversity of stroma was lower than other parts, and the fungal diversity of wild C. sinensis was lower than that of artificial C. sinensis. The β diversity analysis showed that the fungal and bacterial community diversity of soil microhabitat samples was significantly different from that of C. sinensis. The fungal community diversity was less different between wild and artificial C. sinensis, especially in sclerotia. LEfSe analysis showed a lot of species diversity between wild and artificial C. sinensis. Those different species between wild C. sinensis, artificial C. sinensis and their habitat soil provide ideas for further research on breed and components of C. sinensis.


Subject(s)
Cordyceps/genetics , High-Throughput Nucleotide Sequencing , Microbiota/genetics , Soil , Soil Microbiology
12.
Article in Chinese | WPRIM | ID: wpr-887953

ABSTRACT

The plant root-associated microbiomes include root microbiome and rhizosphere microbiome, which are closely related to plant life activities. Nearly 30% of photosynthesis products of plants are used to synthesize root compounds, there is evidence that root compounds regulate and significantly affect the root microbiome Tanshinones are the main hydrophobic components in Salvia miltiorrhiza. In order to study whether these compounds can regulate the root-associated microbiomes of S. miltiorrhiza, our study first identified a white root S. miltiorrhiza(BG) which contains little tanshinones. Retain of the fifth intron of tanshinones synthesis key enzyme gene SmCPS1 leading to the early termination of the SmCPS1 gene, and a stable white root phenotype. Further, wild type(WT) and BG were planted in greenhouse with nutrient soil(Pindstrup, Denmark) and Shandong soil(collected from the S. miltiorrhiza base in Weifang, Shandong), then high-throughput sequencing was used to analyze the root-associated microbiomes. The results showed that the tanshinones significantly affected the root-associated microbiomes of S. miltiorrhiza, and the impact on root microbiomes was more significant. There are significant differences between WT and BG root microbiomes in species richness, dominant strains and co-occurrence network. Tanshinones have a certain repelling effect on Bacilli which belongs to Gram-positive, while specifically attract some Gram-negative bacteria such as Betaproteobacteria and some specific genus of Alphaproteobacteria. This study determined the important role of tanshinones in regulating the structure of root-associated microbiomes from multiple angles, and shed a light for further improving the quality and yield of S. miltiorrhiza through microenvironment regulation.


Subject(s)
Abietanes , Microbiota , Plant Roots , Salvia miltiorrhiza
13.
Acta Pharmaceutica Sinica ; (12): 2295-2301, 2021.
Article in Chinese | WPRIM | ID: wpr-887036

ABSTRACT

Medicinal and edible Armeniacae Semen Amarum (ASA) is susceptible to fungal contamination because it is rich in oil and other nutrients. In this study, the fungal community diversity in ASA samples was analyzed based on a DNA metabarcoding technique to provide evidence for its safe use. Twelve batches of ASA samples samples from four medicinal material markets and three processing approaches were collected. Total DNA was extracted, the ITS2 sequences were amplified, and high-throughput sequencing was performed using the Illumina MiSeq PE300 platform. The results show that Ascomycota was the most dominant fungus in ASA samples. The predominant genus in sample SW1_P was Diutina, whereas the most predominant genus in the other samples was Aspergillus. Three harmful fungi were identified, namely, Aspergillus flavus, Wallemia sebi, and Rhizopus arrhizus. In addition, significant differences were observed in the relative abundance of Botryosphaeriales and Alternaria in ASA samples from different collection sites. Meanwhile, there were significant differences in the relative abundance of Hypocreales and Cladosporium in ASA samples from different processing approaches. In summary, the DNA metabarcoding technique can effectively clarify the fungal community diversity and quickly detect potential toxigenic fungi in ASA samples, thus providing a warning for mycotoxin contamination.

14.
Organ Transplantation ; (6): 403-2021.
Article in Chinese | WPRIM | ID: wpr-881524

ABSTRACT

Objective To evaluate the effect and mechanism of cysteine rich protein 61, namely CCN family member 1(CCN1) on the survival of adipose tissues in rats after autologous fat grafting. Methods At 1 week after the establishment of autologous fat grafting rat models, all animals were randomly divided into the CCN1 group (n=20) and control group (n=20). The survival of fat grafts, the morphology of fat graft tissues, the proportion of active adipocytes and the number of new blood vessels of rats were statistically compared between two groups. The levels of differential expressed messenger ribonucleic acid (mRNA) in the fat graft tissues of rats were compared between two groups by high-throughput sequencing and subsequently subject to cluster analysis. The expression levels of related proinflammatory cytokines of fat graft tissues of rats were statistically compared between two groups. Results The weight retention rate of adipose tissues in the CCN1 group was significantly higher than that in the control group (P < 0.05). In the CCN1 group, the integrity of adipocytes was considerably higher, the degree of vesiculation and vacuolation, the degree of inflammatory cell aggregation and the degree of fibrosis were significantly lower than those in the control group (all P < 0.000 1). Immunofluorescence staining demonstrated that the proportion of active adipocytes with uniform morphology was higher in the CCN1 group, whereas the proportion of active adipocytes was lower and the cells were observed in different sizes accompanied by vesiculation in the control group. Compared with the control group, the quantity of new blood vessels was significantly higher, and the expression levels of platelet derived growth factor (PDGF) and fibroblast growth factor (FGF) mRNA were remarkably higher in the CCN1 group (all P < 0.05). High-throughput sequencing analysis showed that the data at the transcriptome levels significantly differed between two groups. In the CCN1 group, the gene expression levels of cell surface markers, inflammatory cytokines and chemokines related to M1 macrophages tended to decline. Real-time fluorescent quantitative polymerase chain reaction (RT-qPCR) revealed that the mRNA expression levels of interleukin (IL)-8, IL-1 and Toll-like receptor (TLR) 2 in the CCN1 group were significantly lower than those in the control group (P < 0.01-0.05). Conclusions During autologous fat grafting, supplement of exogenous CCN1 may effectively promote the neovascularization of adipose tissues and improve the survival rate of fat graft probably by mediating the transformation of macrophages into M2 phenotype via down-regulating the TLR2 expression level.

15.
Protein & Cell ; (12): 315-330, 2021.
Article in English | WPRIM | ID: wpr-880878

ABSTRACT

Advances in high-throughput sequencing (HTS) have fostered rapid developments in the field of microbiome research, and massive microbiome datasets are now being generated. However, the diversity of software tools and the complexity of analysis pipelines make it difficult to access this field. Here, we systematically summarize the advantages and limitations of microbiome methods. Then, we recommend specific pipelines for amplicon and metagenomic analyses, and describe commonly-used software and databases, to help researchers select the appropriate tools. Furthermore, we introduce statistical and visualization methods suitable for microbiome analysis, including alpha- and beta-diversity, taxonomic composition, difference comparisons, correlation, networks, machine learning, evolution, source tracing, and common visualization styles to help researchers make informed choices. Finally, a step-by-step reproducible analysis guide is introduced. We hope this review will allow researchers to carry out data analysis more effectively and to quickly select the appropriate tools in order to efficiently mine the biological significance behind the data.

16.
Article in Chinese | WPRIM | ID: wpr-862452

ABSTRACT

Objective @# To detect the composition of the subgingival microbiota in generalized aggressive periodontitis (GAgP) and severe chronic periodontitis (SCP) patients tested by high-throughput sequencing (HTS) technologies, analyze its diversity and function by using bioinformatics, and observe changes in the subgingival microbiota before and after periodontal initial therapy.@* Methods@#Eleven patients with GAgP and 14 patients with SCP who visited the Department of Periodontics in Stomatological Hospital of Kunming Medical University from September 2018 to May 2019 were recruited, and subgingival plaque samples were collected at baseline and 6 weeks after initial therapy. Then, the genomic DNA was distracted and sequenced by the Illumina MiSeq high-throughput sequencing platform. QIIME (quantitative insights in microbial ecology), Mothur, SPSS and other software were used to analyze community information. LEfSe difference analysis (linear discriminant analysis effect size), network analysis, and the KEGG PATHWAY database (https://www.kegg.jp/kegg/pathway.html) were used to predict community function. @* Results @# At baseline, the dominant microbiota of GAgP and SCP patients were similar, including Bacteroidetes, Porphyromonas and Porphyromonas endodontalis. Six weeks after initial therapy, as the periodontal pocket became shallower, the variation trend of the microbiota of GAgP and SCP patients was similar. The relative abundance of gram-negative bacteria, such as Bacteroidetes, Porphyromonas and Porphyromonas endodontalis, decreased, while the relative abundance of gram-positive bacteria, such as Proteobacteria, Actinomyces and Rothia aeria, increased. Actinobacteria were significantly increased biomarkers of the subgingival microbiota in GAgP after treatment. Streptococcus is an important genus that connects the microbiota related to periodontitis and the microbiota related to periodontal health. Community function prediction result showed that initial treatment can reduce the functions of amino acid metabolism, methane metabolism, and peptidase in GAgP and SCP patients.@*Conclusion@#The subgingival microbiota of GAgP and SCP patients are similar. Streptococcus, as an early colonizer, may play an important role in promoting plaque biofilm formation and maturation in the process of subgingival flora from health to imbalance. Initial therapy can change the composition and structure of the subgingival microbiota, reduce community diversity, and reduce the functions of amino acid metabolism, methane metabolism, and peptidase in GAgP and SCP patients.

17.
Article in Chinese | WPRIM | ID: wpr-879109

ABSTRACT

The research on endophytes of medicinal plants mainly relies on the traditional culture and isolation methods. Because of their functions such as promoting host growth, improving stress resistance, promoting the accumulation of medicinal active ingredients or directly producing medicinal active ingredients, the endophytes of medicinal plants have gradually attracted wide attention. However, it was found that the strains isolated by traditional methods were not the true dominant endophytes of medicinal plants by comparing the results of traditional culture isolation with high-throughput sequencing. The blind and random nature of traditional methods leads to the lack of standards in terms of medium selection, culture time and interaction between species. On the contrary, high-throughput sequencing technology is an emerging molecular biology technology developed in recent decades. Due to its high resolution level and indepen-dent culture, it can be used for thorough analysis of the community structure and diversity of environmental microorganisms. Therefore, we proposed the strategy of using high-throughput sequencing technology to guide the traditional culture and isolation of endophytes from medicinal plants. Firstly, the endophytic structure and diversity of medicinal plants were completely clear by high-throughput sequencing technology, and the dominant endophytes of the host were unequivocal. Then according to the characteristics of each dominant endophytes design or query suitable medium for its growth to culture and isolation. Finally, the function of the isolates was studied. This method can prevent researchers from missing out on the important functional strains of the host, expand the research scope of endophytes of medicinal plants, and facilitate the in-depth excavation and utilization of endophytes of medicinal plants.


Subject(s)
Endophytes/genetics , High-Throughput Nucleotide Sequencing , Plants, Medicinal , Research Design
18.
China Pharmacy ; (12): 1196-1204, 2021.
Article in Chinese | WPRIM | ID: wpr-876886

ABSTRACT

OBJECTIVE:To study the inhibi tory effects of genistein on the growth of human nasopharyngeal carcinoma. CNE 1 cells and predict its potential target. METHODS :CCK-8 method was used to test the effects of 0(blank control ),12.5,25,50, 100,150 µmol/L genistein on the proliferation of CNE 1 cells after treated for 24,48,72 h. Flow cytometry was carried out to detect the effects of 0(blank control ),15,30,60 µmol/L genistein on the cell cycle and ap optosis of CNE 1 cells after treated for 24 h. Scratch test was used to investigate the effects of 0(blank control ), 10, 20, 30 µmol/L genistein on themigration ability of CNE 1 cells after treated for 24 h. High (No.18210156) throughput sequencing was conducted to discover the differential genes in CNE 1 cells after treated with 0(blankcontrol),30 µmol/L genistein for 24 h. RT-qPCR assay was adopted to verify the mRNA expression of related differential genes in above trials. RESULTS : Compared with blank control,12.5,25,50,100,150 µmol/L genistein sho wed significant inhibitory effect on the proliferation of CNE 1 cells(P< 0.01),in a concentration- time-effect manner ;15,30 µmol/L genistein could arrest CNE 1 cell cycle at G 0/G1 stage(P<0.05 or P< 0.01);30,60 µmol/L could arrest CNE 1 cell cycle at G 2/M stage and promoted cell apoptosis (P<0.05 or P<0.01). 10,20,30 µmol/L genistein could significantly inhibit the migration ability of CNE 1 cells(padj<0.01). High throughput sequencing revealed a total of 2 271 differentialgenes(P<0.05),1 154 of which were up-regulated while 1 117 of which were down-regulated ;8 potential target genes ,including p53,p21,STC2,FGF2,CDK6,CYCLIN D ,PI3K,AKT,were screened by cell experiment. After validated by RT-qPCR assay ,mRNA expression of p53,p21,STC2,FGF2,CDK6,CYCLIN D and AKT were significantly down-regulated(P<0.05),which consistent with the sequencing results. CONCLUSIONS :Genistein can effectively inhibit the growth of human nasopharyngeal carcinoma CNE 1 cells,the mechanism of which may associated with inhibiting the expression of mutant gene p53,restoring the function of wild-type P 53 protein and inhibiting the activity of PI 3K/Akt pathway.

19.
Acta Pharmaceutica Sinica ; (12): 872-878, 2021.
Article in Chinese | WPRIM | ID: wpr-876528

ABSTRACT

In order to provide a scientific basis for the establishment of a Daphnes Cortex medicinal material fungus library and the screening of endophytic fungi that promote the growth of Daphnes Cortex and increase the content of daphnetin, we used Illumina high-throughput testing technology to analyze 9 Daphnes Cortex samples from Gansu and Shanxi provinces. A total of 632 766 valid sequences were obtained, including 348 OTUs, 4 phyla, 20 classes, 48 orders, 108 families, 154 genera, and 208 species. The sum of the first 3 fungal genera account for more than 65% of the total abundance, with the highest reaching 98.4%. Alternaria and Phoma are the main genuses of Daphne giraldii Nitsche, and Altemaria is the dominant genus. The endophytic fungi community of Daphnes Cortex is rich in diversity, and the order of fungal diversity in different producing areas is: Gangu County > Wutai County > Tanchang County.

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Article in Chinese | WPRIM | ID: wpr-875971

ABSTRACT

Objective@# To investigate the influence of mechanical debridement on the subgingival microbiome in chronic periodontitis by 16S rRNA high-throughput sequencing.@*Methods@#Patients with generalized moderate to severe chronic periodontitis were recruited into this study and received oral hygiene instruction and supragingival scaling. One week later, they received ultrasonic and manual subgingival scaling and root planning. Clinical parameters were recorded and subgingival plaques were sampled at baseline and 3 months and 6 months after treatment. The comprehensive profiles of the subgingival microbiome were analyzed by sequencing the V3-4 region of 16S rRNA with the Illumina MiSeq platform.@*Results @#Alpha diversity analysis showed that the richness and diversity of the subgingival community were consistent before and after treatment, but a significant difference in community structure was detected only between baseline and month 3 by principal coordinates analysis (PCoA). After 3 months, the clinical parameter as probing depth (PD) decreased significantly and the relative abundances of the genera related to periodontitis such as Porphyromonas, Treponema, Tannerella, and Filifactor decreased significantly. Meanwhile, the relative abundances of the genera associated with periodontal health increased, such as Capnocytophaga, Kingella. Six months later, however, less genera related to periodontitis decreased significantly from the baseline level, such as Filifactor. PD decreased significantly compared with baseline, but increased significantly compared with 3 months after treatment. @* Conclusion@#Mechanical debridement alone could relieve periodontal inflammation and balance microbial dysbiosis and the greater efficacy occurred 3 months after treatment.

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