For tal gene detection, Xoo genomic DNA was extracted, digested with BamHI, separated in agarose gels, and transferred to membranes for blotting as previously reported [33] (link), [44] (link). The probe was made from a DNA fragment labeled with DIG containing the repetitive region of pthXo1 (GenBank accession number: AY495676). Bacterial Genomic DNA Miniprep Kit was purchased from Axygen (USA). Restriction endonucleases and DNA molecular weight markers were provided by TaKaRa Bio (Japan). DIG-labeled Southern Blot kits were purchased from Roche (Switzerland) and Immobilon-Ny+ membranes were supplied by Millipore (USA).
Bacterial genomic dna miniprep kit
Manufactured by Corning
Sourced in United States
The Bacterial Genomic DNA Miniprep Kit is a laboratory product designed for the rapid and efficient extraction of genomic DNA from bacterial samples. It provides a simple and reliable method for isolating high-quality DNA suitable for various downstream applications, such as PCR, restriction digestion, and sequencing.
Automatically generated - may contain errors
Lab products found in correlation
Immobilon ny membrane, by Merck Group (1 mentions)
Chloramphenicol, by Solarbio (1 mentions)
Kanamycin, by Solarbio (1 mentions)
Sanprep gel extraction kit, by Sangon (1 mentions)
Ampicillin, by Solarbio (1 mentions)
Sanger sequencing, by GenScript (1 mentions)
High fidelity ex taq pcr kit, by Takara Bio (1 mentions)
High fidelity pcr kit, by Takara Bio (1 mentions)
Ta cloning kit, by Takara Bio (1 mentions)
Hiseq 2000 system, by Illumina (1 mentions)
5 protocols using bacterial genomic dna miniprep kit
1
Tal Gene Detection in Xoo
2
Carotenoid production in E. coli
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The strains and plasmids used in this study are listed in Table A1 . E. coli was cultured at 37°C in Lysogeny broth (10 g/L Difco tryptone, 5 g/L Difco yeast extract, and 10 g/L NaCl). The carotenoid fermentation medium was composed of (per liter) 10 g tryptone, 5 g yeast extract, and 10 g NaCl; 2% glycerol (v/v) was added to LB (Lysogeny broth) + glycerol. Apramycin sulfate (50 mg/L; Ruitaibio), chloramphenicol (34 mg/L; Solarbio), ampicillin (100 mg/L; Solarbio), kanamycin (50 mg/L; Solarbio), or β‐D‐1‐thiogalactopyranoside (IPTG, 1 mmol/L; Solarbio) were added to the media, where appropriate. Plasmids were extracted using the Bacterial Genomic DNA Miniprep Kit (Axygen Biosciences). Polymerase chain reaction (PCR) products were digested with DpnI for 0.5 hr at 37°C and then purified using a SanPrep Gel Extraction Kit (Sangon Biotech). Plasmids and PCR products were sequenced using Sanger sequencing (GenScript Co., Ltd).
3
Molecular Serotyping of Pneumococcal Isolates
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The serotypes of 309 S. pneumoniae isolates were identified using PCR as previously reported (Pai et al., 2006 (link); Dias et al., 2007 (link); Da Gloria-Carvalho et al., 2010 (link)). Briefly, genomic DNA of the pneumococcal isolates was extracted using a Bacterial Genomic DNA MiniPrep Kit (Axygen, United States) and then quantified using ultraviolet spectrophotometry (Sun et al., 2010 (link)). Using 100 ng DNA of each of the genomic DNA samples as the template, separate PCRs were performed using a High-Fidelity Ex-Taq PCR Kit (TaKaRa, China) and deferent pairs of serotyping primers (Supplementary Table 1 ). The PCRs were initiated by incubation at 94°C for 5 min for DNA denaturation, followed by 35 cycles at 94°C for 30 s, 52–54°C for 30 s, and 72°C for 30–60 s to amplify the target gene fragments and finally an incubation step at 72°C for 7 min. The S. pneumoniae 16S rRNA gene and cpsA gene, a sequence-conserved capsule formation-determining gene, were used as the controls (El Aila et al., 2010 (link); Coskun-Ari et al., 2012 (link)). For each of the genomic DNA samples, three independent PCR repeats were performed for serotype detection.
4
Leptospiral Genomic DNA Extraction and hslU/hslV Gene Analysis
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Genomic DNA from each of the 12 leptospiral strains was extracted using a Bacterial Genomic DNA Miniprep Kit (Axygen, Union City, CA, USA). By using a High Fidelity PCR Kit (TaKaRa, Dalian, China), the hslU and hslV genes in the DNA samples from the 10 strains of L. interrogans or the two strains of L. biflexa were detected by PCR with the primers hslU1-F/hslU1-R and hslV1-F/hslV1-R or hslU2-F/hslU2-R and hslV2-F/hslV2-R (Table 1 ) because of the large sequence difference at 5′ and 3′ ends of the two genes from L. interrogans and L. biflexa strains. The products were examined on 1.5% ethidium bromide-pre-stained agarose gel after electrophoresis, and then cloned into pMD18-T plasmid to form pMD18-ThslU and pMD18-ThslV using a T-A Cloning Kit (TaKaRa) for sequencing by Invitrogen Co.
5
Bacterial Genome Sequencing Protocol
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Bacterial strains were grown at 37°C in LB broth until the OD600 reached ∼1.0. Cells were harvested by centrifugation for 1 min at 12,000 × g, and genomic DNA was isolated using a bacterial genomic DNA miniprep kit (Axygen, Hangzhou, Zhejiang, China) following the manufacturer’s protocol. The complete genome sequencing procedure was performed using an Illumina HiSeq 2000 system by the Shanghai Haoyu Biotechnology Company. Multiple alignments were performed for the wild-type strain and the derived mutants.
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