Genomic DNAs of IncC-positive strains were extracted by using a MiniBEST Bacteria Genomic DNA Extraction Kit (TaKaRa, Dalian, China), and automatically recovered using the BluePippin (Sage science, United States). The quality and quantity of genomic DNA were confirmed using a Qubit 2.0 fluorometer (Life Technologies). PromethION and Illumina sequencing library preparation were performed using SQK-LSK109 ligation genomic DNA kit (Oxford Nanopore Technologies, United Kingdom) and NEBNext®UltraTM DNA Library Prep Kitfor Illumina (NEB, United States), respectively.
Minibest bacteria genomic dna extraction kit
Manufactured by Takara Bio
Sourced in Japan, China
The MiniBEST Bacteria Genomic DNA Extraction Kit is a lab equipment product designed to extract and purify bacterial genomic DNA. It provides a simple and efficient method for obtaining high-quality DNA from various bacterial species.
Automatically generated - may contain errors
Lab products found in correlation
Minibest whole blood genomic dna extraction kit, by Takara Bio (2 mentions)
Qubit 2.0 fluorometer, by Thermo Fisher Scientific (1 mentions)
Bluepippin, by Sage Science (1 mentions)
Ex taq enzyme, by Takara Bio (1 mentions)
Gel doc xr, by Bio-Rad (1 mentions)
Nextseq 500 platform, by Illumina (1 mentions)
Qiaseq fx dna library kit, by Qiagen (1 mentions)
Qubit dsdna hs assay kit, by Thermo Fisher Scientific (1 mentions)
Labchip gx, by PerkinElmer (1 mentions)
Qiaquick gel extraction kit, by Qiagen (1 mentions)
Phusion high fidelity pcr master mix, by New England Biolabs (1 mentions)
Nanodrop nd 1000 spectrophotometer, by Thermo Fisher Scientific (1 mentions)
Taq pcr master mix, by Vazyme (1 mentions)
Premix ex taq kit, by Takara Bio (1 mentions)
Twistamp basic kit, by Twist Bioscience (1 mentions)
Phosphate buffered saline (pbs), by Solarbio (1 mentions)
Sodium chloride (nacl), by Sinopharm (1 mentions)
Ddh2o, by Solarbio (1 mentions)
Nah2po4, by Sinopharm (1 mentions)
Cycle pure kit, by Omega Bio-Tek (1 mentions)
23 protocols using minibest bacteria genomic dna extraction kit
1
Comprehensive DNA Extraction and Sequencing Protocol
2
Bacterial Genomic DNA Extraction and 16S rRNA Gene Amplification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The bacterial genomic DNA was isolated and purified using the TaKaRa MiniBEST Bacteria Genomic DNA Extraction Kit (Dalian, China). Genomic DNA was then used as the template for PCR amplification of 16S rRNA gene fragments using the bacterial universal primers (27F-5′-AGAGTTGATCCTGGCTCAG-3′ and 1492R-5′-GGTTACCTTGTTACGACTT-3′). The final amplified reaction volume was 50 μl, containing 5.0 μl of 10 × Taq buffers, 4.0 μl of 200 mmol/L dNTPs, 2.0 μl of each primer at 10 μM, 0.5 μl of Ex Taq enzyme (TaKaRa, Dalian), 5.0 μl of genomic DNA, and 31.5 μl of sterilized distilled water. PCR amplification was performed using the Professional Standard 96 Gradient (Biometra, Jena, Germany) with the following cycling parameters: initial denaturation of DNA for 5 min at 95°C, then 30 cycles of denaturation of DNA for 1 min at 94°C, annealing for 1 min at 53°C, extension for 1.5 min at 72°C, and final incubation for 5 min at 72°C (Vasiee et al., 2018 (link)). The PCR products were subsequently purified and sequenced using BGI Biotechnology (Shenzhen, China). DNA sequence alignment was performed using BLAST (https://blast.ncbi.nlm.nih.gov/Blast.cgi ). Finally, phylogenetic trees were constructed using the neighbor-joining (NJ) method implemented in MEGA 5.05 (Arizona State University, Tempe, United States).
3
Analyzing V. parahaemolyticus DNA Fragmentation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
V. parahaemolyticus ATCC 17802 DNA fragmentation following the combination of 405 nm LED treatment and 0.1 mg/mL of citral was analyzed according to the method of Kim et al. [12 (link)] with a slight modification, using the TaKaRa MiniBEST Bacteria Genomic DNA Extraction Kit (TAKARA BIO INC). The purified DNA was dissolved in 150 µL of Elution Buffer. The 40 mL 1% (w/v) agarose gel solution was boiled 3 times, cooled for a moment, and added with 4 µL of GelRed nucleic acid dye. Samples with loading buffer (5:1, v/v) were electrophoresed at 120 V for 30 min. The gel was visualized with GELDOC XR+ (Bio-Rad Laboratories, Co., Ltd., Shanghai, China).
4
Bacterial DNA Extraction for PCR Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA used as template for PCR amplification was isolated from ~0.5–1 mL (1.0–5.0E+09 cells) of a freshly grown (18–24 h) bacterial culture using MiniBEST Bacteria Genomic DNA Extraction Kit (TaKaRa, Japan) according to the manufacturer's instructions. The DNA quantity and purity were measured spectrophotometrically via absorbance measurements at A260 and A280, as well as visualization on 1% agarose gel. A DNA Isolation Reagent for Meat and Meat Products (TaKaRa, Japan) was used to isolate DNA in meat as recommended by the manufacturer.
5
Genomic Analysis of Carbapenem-Resistant Isolates
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The whole genome sequencing of twenty-four carbapenem-resistant isolates was performed with a NextSeq 500 platform (Illumina Inc., San Diego, CA, United States). Briefly, the genomic DNA was extracted using a MiniBEST Bacteria Genomic DNA Extraction Kit (Takara, Dalian, China). To prepare the DNA library for sequencing, a QIAseq FX DNA Library Kits (Qiagen Inc., Valencia, CA, United States) was used following the manufacturer’s recommendations. The quality and quantity of the libraries were assessed with LabChip GX (Perkin Elmer; Waltham, MA, United States) and Qubit dsDNA HS Assay Kit (Life Technologies, United States). All barcoded libraries were pooled together in equimolar amounts and each pool was sequenced on NextSeq 500 in PE-150 bp mode. Later, sequencing raw reads were processed for library adapter removal and filtering using FASTQ preprocessor Fastp v0.12.5 (Chen et al., 2018 (link)) and de novo assembly with SPAdes v3.13.0 (Bankevich et al., 2012 (link)). Genomic repeats were removed from the analyses by filtering out reads that mapped to multiple positions in K. pneumoniae subsp. pneumoniae HS11286 (NCBI accession number: NC_016845 ). Single nucleotide polymorphisms (SNPs) and insertions and deletions (indels) generated by Snippy.
6
Bacterial Genome Sequencing and Bioinformatic Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The genomic DNA was extracted from the strains cultured in NB medium using miniBEST Bacteria Genomic DNA Extraction Kit (TaKaRa code DV810A; TaKaRa, DaLian, China), according to the manufacturer’s instructions, and sent to BGI (Wuhan, China) for quality inspection and genome sequencing. The whole genome sequencing and sequence assembly analysis were performed as described previously [64 (link)].
The raw data were filtered to remove adapters, such as low-quality reads, to generate clean data. The Short Oligonucleotide Analysis Package (SOAPdenovo) software (www.soap.genomics.org.cn ) was used to assemble reads after filtering and perform bioinformatic analysis, including genomic component analysis, comparative genomic analysis, and gene function analysis.
Functional prediction for rRNA, tRNA, and sRNA was performed using the software Glimmer [65 (link)], RNAmmer [66 (link)], tRNAscan [67 (link)], Infernal, and Rfam [68 (link)]. The software Tandem Repeat Finder [69 (link)] was used to predict series repeat sequences, small satellites, and microsatellite sequences; PhiSpy software [70 (link)] was employed to predict the prophage; and CRISPRCas Finder software [71 (link)] was utilized to identify CRISPRs, etc.
The raw data were filtered to remove adapters, such as low-quality reads, to generate clean data. The Short Oligonucleotide Analysis Package (SOAPdenovo) software (
Functional prediction for rRNA, tRNA, and sRNA was performed using the software Glimmer [65 (link)], RNAmmer [66 (link)], tRNAscan [67 (link)], Infernal, and Rfam [68 (link)]. The software Tandem Repeat Finder [69 (link)] was used to predict series repeat sequences, small satellites, and microsatellite sequences; PhiSpy software [70 (link)] was employed to predict the prophage; and CRISPRCas Finder software [71 (link)] was utilized to identify CRISPRs, etc.
7
Bacterial DNA Extraction and 16S rRNA Amplification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bacterial nucleic acids were isolated using the Takara MiniBEST Bacteria Genomic DNA Extraction Kit (Takara, Japan) according to the manufacturer’s protocol, and bacterial DNA extraction was verified by agarose gel electrophoresis.
The V4 hypervariable region of the 16S ribosomal RNA (rRNA) gene was amplified by polymerase chain reaction (PCR) using a specific primer (515F-806R) with a specific barcode. All PCR runs in the present research were conducted with Phusion® High-Fidelity PCR Master Mix (New England Biolabs). Furthermore, the PCR products were mixed with equal volumes of loading buffer (1 × , containing SYBR green) and then electrophoretically separated on a 2% agarose gel. Only the samples with bright main strip bands between 400 and 450 bp could be used for further experimental study. Subsequently, PCR products were further purified using the QIAquick Gel extraction kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions.
The V4 hypervariable region of the 16S ribosomal RNA (rRNA) gene was amplified by polymerase chain reaction (PCR) using a specific primer (515F-806R) with a specific barcode. All PCR runs in the present research were conducted with Phusion® High-Fidelity PCR Master Mix (New England Biolabs). Furthermore, the PCR products were mixed with equal volumes of loading buffer (1 × , containing SYBR green) and then electrophoretically separated on a 2% agarose gel. Only the samples with bright main strip bands between 400 and 450 bp could be used for further experimental study. Subsequently, PCR products were further purified using the QIAquick Gel extraction kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions.
8
Bacterial Genomic DNA Extraction
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Four clinical bacterial strains—S. aureus, Escherichia coli (E. coli), Acinetobacter baumannii (A. baumannii), and Pseudomonas aeruginosa (P. aeruginosa)—were obtained from the Department of Laboratory Medicine, Southwest Hospital (Chongqing, China). The strains were inoculated on blood agar plates overnight at 37 °C in an atmosphere containing 5% CO2. Following incubation, single colonies were lifted off the plates and suspended in sterilized saline water to an optical density of 1.0 at 600 nm. A TaKaRa Mini BEST Bacteria Genomic DNA Extraction Kit was used to extract the bacterial genomic DNA, which were then fragmented by digestion using QuickCut BamHI, EcoRI, and HindIII in 50 μL of reaction mixture at 37 °C for 10 min. Finally, the genomic DNA were diluted to the desired concentration with deionized water. An approximate range from 3–8 μg of genomic DNA can be extracted from 2.0 × 109 of S. aureus cells according to the product manual.
9
Sensitive Detection of Campylobacter jejuni
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The genomic DNA used as a template was extracted from a freshly grown bacterial culture following the manufacturer's protocol with a Takara MiniBEST Bacteria Genomic DNA Extraction Kit. 60 raw chicken meat samples including hearts, thighs, skin samples, and wings that were randomly purchased from the traditional market were used. Approximately 2 g chicken meat samples were added to 18 mL of Brucella enrichment broth followed by incubation at 42°C in anaerobic incubators for 30 h. The bacteria were collected by centrifugation at 8000 g for 5 min for subsequent genomic DNA extraction. The quantity and quality of DNA were tested using a NanoDrop spectrophotometer ND-1000 (NanoDrop Technologies, Wilmington, DE). To assess the sensitivity of the LAMP and PCR assays, 10-fold dilution series of gDNA from C. jejuni (ATCC33252) starting from 34.2 × 100 ng/μL to 34.2 × 10−8 ng/μL was used in the amplification reactions. Furthermore, the DNA from 74 tested isolates used to evaluate the analytical specificity is listed in Table 1 .
10
Quantification of Bacterial DNA Extraction
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Staphylococcus aureus, Streptococcus vestibularis, and Escherichia coli were selected as representatives of each group for quantification. They were procured from Enzynomics (Daejeon, South Korea), National Culture Collection for Pathogens (NCCP, South Korea), and Korean Collection for Type Cultures (KCTC, South Korea), respectively. For the standard control, E. coli and S. aureus were cultured in Luria–Bertani (LB) broth (LPS solution, Daejeon, South Korea), and S. vestibularis was cultured in tryptic soy broth (MBcell, Seoul, South Korea) at 37°C. After culturing, 100 μL of bacteria was diluted (1:10) in culture media and smeared on agar plates. Further, 100 μL of the cultured bacteria was used for DNA extraction, which was performed using the TaKaRa MiniBEST Bacteria Genomic DNA Extraction Kit (TaKaRa, Tokyo, Japan) according to the manufacturer’s protocol. The extracted genomic DNA was eluted in 100 μL nuclease-free water. A standard curve of DNA/colony-forming unit (CFU) was calculated using comparative analysis of the number of cultured colonies.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!