Microbial Diversity and Physicochemical Characteristics of the Maotai-Flavored Liquor Fermentation Process

2020 ◽  
Vol 20 (7) ◽  
pp. 4097-4109 ◽  
Author(s):  
Yijie Dai ◽  
Zhiqiang Tian ◽  
Wangni Meng ◽  
Zongjun Li
Keyword(s):  
Amylase Activity ◽  
Rna Extraction ◽  
Physicochemical Characteristics ◽  
Molecular Techniques ◽  
Its Sequencing ◽  
Total Rna ◽  
Microbial Community Structures ◽  
Rrna Sequencing ◽  
Fermented Grains ◽  
Dominant Bacteria

The grains fermented in Maotai-flavored liquor can be classified into two groups: high-temperature fermented grains generated by stacking and grains fermented in pits. The Maotai-flavor making process enriches a special microbiota from the fermented grains, which makes related studies more difficult. The use of modern molecular techniques to detect and analyze diversity and changes in total bacterial (16S rRNA sequencing) and fungal (internal transcribed spacer (ITS) sequencing) counts have helped to overcome the shortcomings of traditional technological research. The total RNA extracted by Fe3O4–SiO2 nanoparticles was retrieved into DNA by MagBeads Total RNA Extraction Kit. However, discrepancies exist in the microbial community structures at different fermentation periods. Dominant bacteria include Escherichia-Shigella, Lactobacillus, Clostridium, and Streptococcus species and dominant fungi include Alternaria, Ciliophora, Pyrenochaetopsis, Cyphellophora, Aspergillus, Issatchenkia, Pichia, Candida, and Zygosaccharomyces species. Analyses of the enzymatic activity of samples at different fermentation stages have revealed that some existing bacilli influence amylase activity. Here, to investigate flavor changes during each fermentation round, the liquor quality at all rounds was comprehensively assessed based on the corresponding physicochemical properties, which were summarized by sensory evaluation. These results suggested that the liquor quality in the third, fourth, and fifth fermentation rounds was superior.

scholarly journals High-quality total RNA isolation from melon (Cucumis melo L.) fruits rich in polysaccharides

2017 ◽  
Vol 38 (4) ◽  
pp. 2201 ◽  
Author(s):  
Gabrielle Silveira de Campos ◽  
Ricardo Antônio Ayub ◽  
Rafael Mazer Etto ◽  
Carolina Weigert Galvão ◽  
Marília Aparecida Stroka ◽  
...  
Keyword(s):  
Rna Extraction ◽  
Sodium Perchlorate ◽  
Rna Isolation ◽  
World Market ◽  
Molecular Techniques ◽  
Guanidine Thiocyanate ◽  
High Quality ◽  
Total Rna ◽  
High Concentrations ◽  
Cucumis Melo L

Melon, a member of the family Cucurbitaceae, is the fourth most important fruit in the world market and, on a volume basis, is Brazil’s main fresh fruit export. Many molecular techniques used to understand the maturation of these fruits require high concentrations of highly purified RNA. However, melons are rich in polyphenolic compounds and polysaccharides, which interfere with RNA extraction. This study aimed to determine the most appropriate method for total RNA extraction from melon fruits. Six extraction buffers were tested: T1) guanidine thiocyanate/phenol/chloroform; T2) sodium azide/?-mercaptoethanol; T3) phenol/guanidine thiocyanate; T4) CTAB/PVP/?-mercaptoethanol; T5) SDS/sodium perchlorate/PVP/?-mercaptoethanol, and T6) sarkosyl/PVP/guanidine thiocyanate, using the AxyPrepTM Multisource Total RNA Miniprep Kit. The best method for extracting RNA from both mature and green fruit was based on the SDS/PVP/?-mercaptoethanol buffer, because it rapidly generated a high quality and quantity of material. In general, higher amounts of RNA were obtained from green than mature fruits, probably due to the lower concentration of polysaccharides and water. The purified material can be used as a template in molecular techniques, such as microarrays, RT-PCR, and in the construction of cDNA and RNA-seq data.

scholarly journals THE MODULATION OF DRUG EFFLUX TRANSPORTER BY CURCUMIN IN MCF7 BREAST CANCER CELLS AFTER REPEATED EXPOSURE OF ENDOXIFEN AND ESTRADIOL

2018 ◽  
Vol 10 (1) ◽  
pp. 102
Author(s):  
Robby Hertanto ◽  
Wilson Bastian ◽  
Paramita . ◽  
Melva Louisa
Keyword(s):  
Breast Cancer ◽  
Cell Viability ◽  
Mrna Expression ◽  
Rna Extraction ◽  
Efflux Transporters ◽  
Drug Efflux ◽  
Mcf7 Cells ◽  
Total Rna ◽  
P Glycoprotein ◽  
Drug Efflux Transporters

Objective: The aim of the present study was to determine whether curcumin (CM) can prevent drug sensitivity of breast cancer (BC) cells when E andβ-E2 are administered together and whether the underlying mechanism involves modulation of drug efflux transporters.Methods: MCF7 BC cells were treated with the vehicle only, E+β-E2, or E+β-E2+CM repeatedly for 8 weeks. Afterward, the cells were harvested,counted, and isolated for total RNA extraction. Total RNA was then processed into cDNA and further processed for the determination of mRNAexpression patterns of drug efflux transporters (P-glycoprotein, BCRP, and MRP1).Results: Decreased sensitivity of BC cells was shown by the increased cell viability of MCF7 cells after 8 weeks. This condition was accompanied withincreased mRNA expression of P-glycoprotein, BCRP, and MRP1 in cells treated with E+β-E2, as compared with the vehicle only. CM, administered incombination with E+β-E2, resulted in decreased cell viability versus E and β-E2 and also decreased in mRNA expression of P-glycoprotein, BCRP, andMRP1.Conclusion: CM partially reversed the sensitivity loss of BC cells to E in the presence of β-E2 by modulating drug efflux transporters.

scholarly journals Clinical optochin resistant Streptococcus pneumoniae and Streptococcus pseudopneumoniae strains in Tunisia

2021 ◽  
Vol 15 (05) ◽  
pp. 672-677
Author(s):  
Sonia Ktari ◽  
Nour El Houda Ben Ayed ◽  
Sonda Maalej ◽  
Basma Mnif ◽  
Faouzia Rhimi ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
16S Rrna ◽  
Molecular Techniques ◽  
16S Rrna Sequencing ◽  
Easy Method ◽  
Rrna Sequencing ◽  
C Subunit ◽  
Density Values ◽  
A Subunit ◽  
Human Infections

Introduction: Streptococcus pneumoniae can be responsible for severe human infections. Optochin resistance has been a potential cause of misidentification of pneumococcus and other members of the mitis group. Hence, rapid and easy optochin resistant (Optr) S. pneumoniae identification is essential. Methodology: Atypical pneumococci were characterized using optochin susceptibility, bile solubility based on spectrophotometric reading, serotyping, pulsed field gel electrophoresis (PFGE), 16S rRNA sequencing and PCR-based assays targeting pneumococcal genes lytA, ply, pspA, cpsA, Spn9802 and Spn9828. Results: Optical density values for the bile solubility test suggest the identification of four Optr S. pneumoniae and one Streptococcus pseudopneumoniae. All Optr pneumococci harbored cpsA, lytA, ply, Spn9802, Spn9828 and pspA genes. Only ply, spn9802 and Spn9828 genes were detected in S. pseudopneumoniae. The 16S rRNA sequencing differentiates between these two species. Optr S. pneumoniae strains belonged to different genotypes and serotypes (14, 19A, 3 and 9V). Three Optr S. pneumoniae isolates were typed as pspA family 2, while one belonged to pspA family 1. Sequencing of the atpA and atpC gene of the Optr variants revealed three mutations in the ATPase a-subunit (L99I, M23V and V52I) and one mutation in ATPase c-subunit (V48I). Conclusions: Our data indicate that bile OD-values provides an accurate, fast and easy method to discriminate between Optr S. pneumoniae and other Streptococcus mitis group. Moreover molecular techniques, confirming the bile test, can be used in order to prevent these atypical pneumococci and alert clinical microbiologists of the presence of these strains in the community.

Isolation of high-quality total RNA from rumen anaerobic bacteria and fungi, and subsequent detection of glycoside hydrolases

2011 ◽  
Vol 57 (7) ◽  
pp. 590-598 ◽  
Author(s):  
Pan Wang ◽  
Meng Qi ◽  
Perry Barboza ◽  
Mary Beth Leigh ◽  
Emilio Ungerfeld ◽  
...  
Keyword(s):  
Gene Expression ◽  
Solid Phase ◽  
Large Subunit ◽  
Rna Extraction ◽  
Small Subunit ◽  
Glycoside Hydrolases ◽  
Small Subunit Rrna ◽  
High Quality ◽  
Major Barrier ◽  
Total Rna

The rumen is one of the most powerful fibrolytic fermentation systems known. Gene expression analyses, such as reverse transcription PCR (RT-PCR), microarrays, and metatranscriptomics, are techniques that could significantly expand our understanding of this ecosystem. The ability to isolate and stabilize representative RNA samples is critical to obtaining reliable results with these procedures. In this study, we successfully isolated high-quality total RNA from the solid phase of ruminal contents by using an improved RNA extraction method. This method is based on liquid nitrogen grinding of whole ruminal solids without microbial detachment and acid guanidinium – phenol – chloroform extraction combined with column purification. Yields of total RNA were as high as 150 µg per g of fresh ruminal content. The typical large subunit/small subunit rRNA ratio ranged from 1.8 to 2.0 with an RNA integrity number (Agilent Technologies) greater than 8.5. By eliminating the detachment step, the resulting RNA was more representative of the complete ecosystem. Our improved method removed a major barrier limiting analysis of rumen microbial function from a gene expression perspective. The polyA-tailed eukaryotic mRNAs obtained have successfully been applied to next-generation sequencing, and metatranscriptomic analysis of the solid fraction of rumen contents revealed abundant sequences related to rumen fungi.

scholarly journals Comparative analysis of gut microbiota among the male, female and pregnant giant pandas (Ailuropoda Melanoleuca)

2019 ◽  
Vol 14 (1) ◽  
pp. 288-298
Author(s):  
Siyue Zhao ◽  
Caiwu Li ◽  
Guo Li ◽  
Shengzhi Yang ◽  
Yingming Zhou ◽  
...  
Keyword(s):  
Comparative Analysis ◽  
16S Rrna ◽  
Gut Microbiota ◽  
High Throughput Sequencing ◽  
Vital Role ◽  
Ailuropoda Melanoleuca ◽  
Its Sequencing ◽  
Giant Pandas ◽  
Host Health ◽  
Rrna Sequencing

AbstractThe giant panda (GP) was the most endangered species in China, and gut microbiota plays a vital role in host health. To determine the differences of the gut microbiota among the male, female and pregnant GPs, a comparative analysis of gut microbiota in GPs was carried out by 16S rRNA and ITS high-throughput sequencing. In 16S rRNA sequencing, 435 OTUs, 17 phyla and 182 genera were totally detected. Firmicutes (53.6%) was the predominant phylum followed by Proteobacteria (37.8%) and Fusobacteria (7.1%). Escherichia/Shigella (35.9%) was the most prevalent genus followed by Streptococcus (25.9%) and Clostridium (11.1%). In ITS sequencing, 920 OTUs, 6 phyla and 322 genera were also detected. Ascomycota (71.3%) was the predominant phylum followed by Basidiomycota (28.4%) and Zygomycota (0.15%). Purpureocillium (4.4%) was the most prevalent genus followed by Cladosporium (2.5%) and Pezicula (2.4%). Comparative analysis indicated that the male GPs harbor a higher abundance of phylum Firmicutes than female GPs with the contribution from genus Streptococcus. Meanwhile, the female GPs harbor a higher abundance of phylum Proteobacteria than male GPs with the contribution from genus Escherichia/ Shigella. In addition, the shift in bacteria from female to pregnant GPs indicated that phylum Firmicutes increased significantly with the contribution from Clostridium in the gut, which may provide an opportunity to study possible associations with low reproduction of the GPs.

Total RNA Extraction from Caenorhabditis elegans

2020 ◽  
Vol 2020 (9) ◽  
pp. pdb.prot101683
Author(s):  
Michael R. Green ◽  
Joseph Sambrook
Keyword(s):  
Caenorhabditis Elegans ◽  
Rna Extraction ◽  
Total Rna ◽  
Total Rna Extraction

A RAPID AND EFFECTIVE METHOD FOR TOTAL RNA EXTRACTION FROM CONIDIA AND MYCELIUM OF SEPTORIA TRITICI

2003 ◽  
Vol 11 (1) ◽  
pp. 53-59 ◽  
Author(s):  
JIAN-RONG GUO ◽  
KAMEL A. ABD-ELSALAM ◽  
FRANK SCHNIEDER ◽  
JOSEPH-ALEXANDER VERREET
Keyword(s):  
Rna Extraction ◽  
Septoria Tritici ◽  
Total Rna ◽  
Total Rna Extraction

scholarly journals Detection of Viable Cryptosporidium parvum in Soil by Reverse Transcription–Real-Time PCR Targetinghsp70mRNA

2011 ◽  
Vol 77 (18) ◽  
pp. 6476-6485 ◽  
Author(s):  
Zhanbei Liang ◽  
Ann Keeley
Keyword(s):  
Cryptosporidium Parvum ◽  
Reverse Transcription ◽  
Rna Binding ◽  
Direct Detection ◽  
Rna Extraction ◽  
Milk Powder ◽  
Extraction Methods ◽  
Content Type ◽  
Total Rna ◽  
Rna Extraction Methods

ABSTRACTExtraction of high-quality mRNA fromCryptosporidium parvumis a key step in PCR detection of viable oocysts in environmental samples. Current methods for monitoring oocysts are limited to water samples; therefore, the goal of this study was to develop a rapid and sensitive procedure forCryptosporidiumdetection in soil samples. The efficiencies of five RNA extraction methods were compared (mRNA extraction with the Dynabeads mRNA Direct kit after chemical and physical sample treatments, and total RNA extraction methods using the FastRNA Pro Soil-Direct, PowerSoil Total RNA, E.Z.N.A. soil RNA, and Norgen soil RNA purification kits) for the direct detection ofCryptosporidiumwith oocyst-spiked sandy, loamy, and clay soils by using TaqMan reverse transcription-PCR. The study also evaluated the presence of inhibitors by synthesis and incorporation of an internal positive control (IPC) RNA into reverse transcription amplifications, used different facilitators (bovine serum albumin, yeast RNA, salmon DNA, skim milk powder, casein, polyvinylpyrrolidone, sodium hexametaphosphate, andSalmonella entericaserovar Typhi) to mitigate RNA binding on soil components, and applied various treatments (β-mercaptoethanol and bead beating) to inactivate RNase and ensure the complete lysis of oocysts. The results of spiking studies showed thatSalmonellacells most efficiently relieved binding of RNA. With the inclusion ofSalmonelladuring extraction, the most efficient mRNA method was Dynabeads, with a detection limit of 6 × 102oocysts g−1of sandy soil. The most efficient total RNA method was PowerSoil, with detection limits of 1.5 × 102, 1.5 × 103, and 1.5 × 104C. parvumoocysts g−1soil for sandy, loamy, and clay samples, respectively.

198 PROGESTERONE, ESTROGEN (ER-α AND ER-β), AND OXYTOCIN RECEPTOR GENE EXPRESSION IN CANINE EMBRYOS

2013 ◽  
Vol 25 (1) ◽  
pp. 248
Author(s):  
A. A. P. Derussi ◽  
A. C. S. Castilho ◽  
R. W. A. Souza ◽  
R. Volpato ◽  
C. R. F. Guaitolini ◽  
...  
Keyword(s):  
Gene Expression ◽  
Relative Abundance ◽  
Hormone Receptors ◽  
Target Genes ◽  
Receptor Gene ◽  
Rna Extraction ◽  
Amplification Efficiency ◽  
Mrna Levels ◽  
Lh Surge ◽  
Total Rna

The aim of this study was to compare the mRNA levels of hormone receptor for progesterone (PR), oestrogen α (ER-α), oestrogen β (ER-β), and oxytocin (OTR) in canine morulae and blastocysts. Ten healthy mature bitches were inseminated based on monitoring vaginal cytology and progesterone concentration. The first insemination was performed on Day 2 after the preovulatory LH surge (progesterone 4 ng mL–1), and the second was performed 48 h later. All females were submitted to ovariohysterectomy (OVH), and the oviduct as well the uterurs were flushed with PBS solution to obtain the embryos. The females were divided into two groups: Group A (n = 5), morulae were collected 8 days after the LH surge and Group B (n = 5), blastocysts were collected 12 days after the LH surge. The pools (n = 10) of embryos (5 embryos/pool) were stored in RNAlater® (Ambion, Life Technologies, USA) at –80°C. The samples were analysed together. The RNA later was removed used PBS calcium free and the total RNA extraction was performed using the Qiagen RNeasy micro-kit (Hildesheim, Germany). Before reverse-transcription (RT) reaction, the total RNA was treated with DNase I Amplification Grade (Invitrogen Life Technologies, Carlsbad, CA, USA). The gene expression of target genes was assessed by real-time RT-qPCR, using SuperScript III for RT and power SYBR Green PCR Master Mix (Applied Biosystems, USA) for cDNA for PCR. The primers for target genes were designed using the software Primer Express® (Applied Biosystems, USA). The gene expression of target genes was normalized by HPRT gene and the relative abundance of mRNA was determined by the ΔΔct method corrected by amplification efficiency using Pffafl’s equation. The means of mRNA relative abundance were compared by t-test. The PR mRNA expression only in blastocysts is similar to the results obtained by Hou et al. (1997) in rat embryos. It is believed that the absence of PR in the early stages of cleavage is due to the indirect action of progesterone by growth factors produced by the maternal reproductive tract (2). Apparently, ER-β action does not occur in the embryo canine phases analysed; however, the action of ER-α seems related to the deployment signal as seen by Hou et al. (1996) in rats. Similarly to findings in the literature, OTR expression decreased in canine embryonic development. This receptor was produced by blastocysts while present in the uterus, which may represent an incidental mechanism to the embryo control of endometrial receptivity, such as also to prevent the development of endometrial luteolytic mechanism. The variation in hormone receptors gene expression in canine embryos can be influencing the transition from morula to blastocyst. In addition, a hormonal influence on these structures can occur in different ways.

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