DNA was isolated from frozen tissue using Qiagen Genomic Tip 500/G (Qiagen, Germantown, MD) or Roche DNA Isolation Kit for Cells and Tissues (Roche, Indianapolis, IN) as described by the manufacturer. One hundred µg of DNA in a final volume of 450 µl water with 10 mM MgCl2 was submitted to a series of enzymatic steps to prepare individual deoxynucleotides: 1) 375 units Dnase I (Life Technologies, Carlsbad, CA) at 37°C for 1.5 hours; 2) 7 units nuclease P1 (Sigma-Alrich, St. Louis, MO) at 37°C for 1.5 hours; 3) 30 units shrimp alkaline phosphatase (Thermo Scientific, Waltham, MA) at 37°C for 1 hours. After DNA digestion, a 50 µl aliquot was set aside for HPLC base analysis. Forty pg of 15N-labeled (±)-anti-cis-DBCDE-dA and (±)-anti-trans-DBCDE-dA was spiked into each sample to serve as an internal standard. One ml ethanol was added to each sample and the sample was incubated at −20°C overnight. The following day the samples were centrifuged at 14,000g for 10 minutes at 4°C. The supernatant was then transferred to a new 1.7 ml microcentrifuge tube and evaporated under a nitrogen stream to concentrate the sample to 50 µl. Fifty µl of methanol was added to bring the samples to 100 µl (1 µg DNA/µl). The samples were filtered through a costar-X spin tube before being transferred to an autosampler vial.
Dna isolation kit for cells and tissue
Manufactured by Roche
Sourced in Germany, United States
The DNA Isolation Kit for Cells and Tissues is a product designed to extract and purify DNA from a variety of cell and tissue samples. The kit provides a reliable and efficient method for obtaining high-quality genomic DNA, which is essential for various downstream applications, such as PCR, sequencing, and molecular analysis.
Automatically generated - may contain errors
Lab products found in correlation
Nanodrop 8000 uv vis spectrophotometer, by Thermo Fisher Scientific (2 mentions)
Shrimp alkaline phosphatase, by Thermo Fisher Scientific (1 mentions)
Dnase 1, by Thermo Fisher Scientific (1 mentions)
Nuclease p1, by Merck Group (1 mentions)
Genomic tip 500 g, by Qiagen (1 mentions)
Abi prism 3700, by Thermo Fisher Scientific (1 mentions)
Bigdye terminator sequencing kit, by Thermo Fisher Scientific (1 mentions)
Onestep qmethyl kit, by Zymo Research (1 mentions)
Dnazap, by Thermo Fisher Scientific (1 mentions)
Phosphodiesterase 1, by Merck Group (1 mentions)
Dnase 1, by Takara Bio (1 mentions)
Ods reverse phase column, by Phenomenex (1 mentions)
Hoescht 3342, by Thermo Fisher Scientific (1 mentions)
Dm500, by Leica camera (1 mentions)
Epoch 2 microplate spectrophotometer, by Agilent Technologies (1 mentions)
Genomestudio methylation module software, by Illumina (1 mentions)
Nanodrop nd 1000 spectrometer, by Thermo Fisher Scientific (1 mentions)
Dimethylmethylene blue, by Merck Group (1 mentions)
Quant it picogreen dsdna reagent and kit, by Thermo Fisher Scientific (1 mentions)
Proteinase k, by Merck Group (1 mentions)
20 protocols using dna isolation kit for cells and tissue
1
DNA Isolation and Deoxynucleotide Preparation
2
Aedes albopictus Genomic Diversity Across China
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Aedes albopictus were collected from 12 sites on public land in Guangzhou, China, between September 23rd and October 22nd, 2015. The sample sites were distributed across an area of 380 km2 through nine administrative districts, and were separated by distances ranging from 2.73 km to 49.34 km (Fig 1 ). All subsequent references to sites follow the numerical designations in Fig 1 . At each site, natural containers were searched for larvae and pupae that were collected and then combined into a single sample to be reared in the laboratory until eclosion.
Samples from Sites 1–8 were raised in tap water containing shrimp powder, while samples from Sites 9–12 were raised in tap water containing shrimp powder, bovine liver powder and nutritional yeast. Following eclosion, adults were fed on 10% sugar solution and allowed to mate, but not bloodfeed. After 4–6 days, 20 females from each sample were killed by freezing and stored in ethanol until DNA extraction. Genomic DNA was extracted using Roche DNA Isolation Kit for Cells and Tissues (Roche, Pleasanton, CA, USA), with an additional step of RNAse treatment. From the 12 sample sites, we selected 152 individuals for ddRAD sequencing.
Samples from Sites 1–8 were raised in tap water containing shrimp powder, while samples from Sites 9–12 were raised in tap water containing shrimp powder, bovine liver powder and nutritional yeast. Following eclosion, adults were fed on 10% sugar solution and allowed to mate, but not bloodfeed. After 4–6 days, 20 females from each sample were killed by freezing and stored in ethanol until DNA extraction. Genomic DNA was extracted using Roche DNA Isolation Kit for Cells and Tissues (Roche, Pleasanton, CA, USA), with an additional step of RNAse treatment. From the 12 sample sites, we selected 152 individuals for ddRAD sequencing.
3
Detecting H. pylori in Vegetables
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA from 1 mL of each vegetable and salad sample was extracted by a DNA isolation kit for cells and tissues (Roche Applied Science, Germany, 11814770001), according to the manufacturer’s instructions. Extracted genomic DNA was amplified for the ureC gene and detected with specific primers HP-F: 5'-GAATAAGCTTTTAGGGGTGTTAGGGG-3’, HP-R: 5'GCTTACTTTCTAACACTAACGCGC-3'. The gene product was 294 bp. The PCR conditions and temperatures were based on the Rahimi and Kheirabadi protocol (10 (link)). Samples inoculated with H. pylori were used as positive controls.
4
Isolation and Characterization of Endophytic Bacteria
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
In order to isolate endophytes, approximately 2 mm of surface plant tissues including the cuticle were discarded under sterile conditions. The remaining plant material was macerated in a sterile mortar and resuspended in 10 mM MgSO4 (1:10 w:v). Epiphytic suspensions and soil dilutions in 10 mM MgSO4 were inoculated on plates (1.6% agar) of methanol mineral salts medium (MMSM; 21) containing 0.5% methanol; 6.89 mM K2HPO4; 4.56 mM KH2PO4; 0.228 mM CaCl2; 0.811 mM MgSO4; 1.71 mM NaCl; 3.7 μM FeCl3; 3.8 mM (NH4)2SO4; 20 nM CuSO4; 41.5 nM MnSO4; 38 nM Na2MoO4; 0.163 μM H3BO3; 0.243 μM ZnSO4; and 21 nM CoCl2, and incubated at 30°C for 8–10 d. Isolated bacterial colonies were streaked in the same medium and incubated at 30°C until growth was observed. Isolated colonies were grown in the same medium and also in GP containing (L−1): Casein peptone 10 g, glycerol 10 g, and agar 15 g. DNA was extracted from cells growing in MMSM medium with the DNA Isolation Kit for Cells and Tissues (Roche Diagnostics, Indianapolis, IN, USA) following the recommended instructions of the supplier.
5
Mutation Identification in Sanjad-Sakati Syndrome
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Genomic DNA was extracted from peripheral blood samples of each patient and their parents using the salting out method. In those receiving prenatal testing, genomic DNA was obtained from 30 mg of chorionic villi at 13 weeks of pregnancy using a DNA isolation kit for cells and tissues (Roche). In order to identify the mutation in 29 SSS patients, polymerase chain reaction (PCR) was performed for exon 3 of the TBCE gene using primers and conditions that were previously described.1 (link) After that, PCR products were subjected to Sanger sequencing using the ABI Prism 3700 apparatus (BigDye Terminator sequencing kit, Applied Biosystems), and sequencing data were compared with the reference sequence (NM_001287801). If a mutation was detected, parents of that patient were analyzed for the validation of segregation, and prenatal testing was carried out for eight families by also using Sanger sequencing.
6
Quantitative DNA Methylation Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA was isolated from the established cell cultures (5 x 106 cells
approximately) using DNA Isolation Kit for Cells and Tissues (Roche Diagnostics,
Mannheim, Germany). Methylation analyses were performed with One-Step qMethyl Kit
(Zymo Research, Irvine, CA, USA) using primers amplifying the MEFVCpG island, analyzed via qRT-PCR. Two reactions were setup as Test and Reference
reactions: The Test reaction includes Methylation Sensitive Restriction Enzymes
(MSREs) to cut at the methylated nucleotides, whereas the Reference reaction does not
contain these enzymes. Therefore, the Test reaction samples are cut if methylated,
creating smaller fragments, which result in lower Ct values.
The data was analyzed with qMethyl Calculator, which calculates the methylation ratio
as follows: Percent methylation = 100 x 2-ΔCt.
where ΔCt is the average Ct value from the Test reaction minus the average Ct value
from the Reference reaction.
approximately) using DNA Isolation Kit for Cells and Tissues (Roche Diagnostics,
Mannheim, Germany). Methylation analyses were performed with One-Step qMethyl Kit
(Zymo Research, Irvine, CA, USA) using primers amplifying the MEFVCpG island, analyzed via qRT-PCR. Two reactions were setup as Test and Reference
reactions: The Test reaction includes Methylation Sensitive Restriction Enzymes
(MSREs) to cut at the methylated nucleotides, whereas the Reference reaction does not
contain these enzymes. Therefore, the Test reaction samples are cut if methylated,
creating smaller fragments, which result in lower Ct values.
The data was analyzed with qMethyl Calculator, which calculates the methylation ratio
as follows: Percent methylation = 100 x 2-ΔCt.
where ΔCt is the average Ct value from the Test reaction minus the average Ct value
from the Reference reaction.
7
Genomic DNA Isolation from HNSCC
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Genomic DNA was isolated from all HNSCC and healthy tissues using the DNA Isolation kit for cells and tissues (catalog no. 11814770001; Roche Diagnostics, Ltd.) following the manufacturer's instructions. DNA concentration and quality were measured with the NanoDrop 8000 UV–Vis Spectrophotometer (Thermo Fisher Scientific, Inc.). DNA sample preparation and hybridization to oligonucleotide arrays was carried out at the Head and Neck Cancer Research laboratory, Johns Hopkins School of Medicine.
8
Genomic DNA Extraction from Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Genomic DNA (gDNA) was extracted from ≈100 mg of tissue using the DNA Isolation Kit for Cells and Tissues (Roche Applied Science, Indianapolis, IN) following the manufacturer's instructions. For quality control purposes, β-actin gene PCR amplification was used to assess gDNA integrity; samples from which β-actin could not be amplified were excluded from the study.
The following standardized precautions were taken to minimize sample-to-sample cross-contamination: all instruments and work benches were wiped down with DNAZap (Ambion, Foster City, CA), followed by 10% bleach and 70% ETOH prior to sample manipulation. New, sterile blades were used for each sample. Tissue processing and nucleotide extraction were limited to a maximum of 10 samples per day. Approximately 100 mg of each specimen was randomly coded in a blinded manner and used for further analyses.
The following standardized precautions were taken to minimize sample-to-sample cross-contamination: all instruments and work benches were wiped down with DNAZap (Ambion, Foster City, CA), followed by 10% bleach and 70% ETOH prior to sample manipulation. New, sterile blades were used for each sample. Tissue processing and nucleotide extraction were limited to a maximum of 10 samples per day. Approximately 100 mg of each specimen was randomly coded in a blinded manner and used for further analyses.
9
Viral Detection and Antibody Determination
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For viral detection and determination of specific antibodies, we used blood, skin biopsy, and throat swab samples from patients. After patient consent forms were obtained, one skin specimen or mucosal specimens were obtained by punch biopsy (3 mm) and blood samples were taken from all patients. In addition, a sample from the throat was collected by swabbing. All samples were transferred in cold chain conditions to the virology laboratory. As soon as samples were received, preparation and storage were carried out. Sera (for serology) and buffy coat (for virus detection) were isolated from whole blood and stored at −20°C. DNA extraction from buffy coat, skin biopsy, and throat swab were performed using DNA Isolation Kit for Cells and Tissues according to the manufacturers’ instructions (Roche, Berlin, Germany). After DNA extraction, it was eluted in 50 μL of buffer and then adjusted to a definitive concentration of 500 ng/μL.
10
Conditional CdGAP Knockout Mice Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
CdGAPflox/flox mice, which have conditional floxed exon 1 allele (C57BL/6 background), systemic CdGAP deficient mice (C57BL/6 background), and Podocin-Cre mice (mixed ICR/129/B6 background) have been described previously27 (link),51 (link). Male mice were used for experiments. Cre-negative mice were used as controls for CdGAPflox/flox;Pod-Cre (CdGAPPod-/-) mice. Spot urine samples were collected at the indicated time points. To determine the genotype, mice DNA was extracted from isolated glomeruli by DNA Isolation Kit for Cells and Tissues (Roche Diagnostics, 11814770001), and subjected to PCR. The genotyping primer sequences used were as follows:
Cre-F 5′-gcttctgtccgtttgccg-3′.
Cre-R 5′-actgtgtccagaccaggc-3′.
CdGAP -F 5′-cctgcgctgtgcaaagagcct-3′.
CdGAP -R 5′-cccaaagtttaagacccgagcctc-3′.
Cre-F 5′-gcttctgtccgtttgccg-3′.
Cre-R 5′-actgtgtccagaccaggc-3′.
CdGAP -F 5′-cctgcgctgtgcaaagagcct-3′.
CdGAP -R 5′-cccaaagtttaagacccgagcctc-3′.
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!