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High capacity dna archive kit

Manufactured by Thermo Fisher Scientific
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The High Capacity DNA Archive Kit is a laboratory product designed for the long-term storage and preservation of DNA samples. It offers high-capacity storage capabilities to maintain the integrity of DNA samples over extended periods.

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Lab products found in correlation

Qiazol reagent, by Qiagen (4 mentions) Dnase 1, by Thermo Fisher Scientific (4 mentions) Rneasy micro kit, by Qiagen (4 mentions) Tri reagent, by Merck Group (2 mentions) Dnaase 1, by Thermo Fisher Scientific (2 mentions) Qiazol lysis reagent, by Qiagen (2 mentions) Primer express software, by Thermo Fisher Scientific (2 mentions) Sequence detection software 1, by Thermo Fisher Scientific (1 mentions) Taqman assay, by Thermo Fisher Scientific (1 mentions) Abi 7500ht machine, by Thermo Fisher Scientific (1 mentions) Sybr green pcr master mix, by Thermo Fisher Scientific (1 mentions) High capacity rna to cdna kit, by Thermo Fisher Scientific (1 mentions) Gapdh, by Thermo Fisher Scientific (1 mentions) Taqman microrna reverse transcription kit, by Thermo Fisher Scientific (1 mentions) Mirvana mirna isolation kit, by Thermo Fisher Scientific (1 mentions) Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions) Primer express, by Thermo Fisher Scientific (1 mentions) Cfx96 touch real time pcr detection system, by Bio-Rad (1 mentions) Sybr green pcr master mix, by Bio-Rad (1 mentions)

13 protocols using high capacity dna archive kit

1

Radiation-Induced Gene Expression Analysis

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Consenting donors or volunteers from the local ethnic population group of Shillong have been recruited in the study, which is ongoing. Withdrawal of blood, irradiation, postirradiation incubation, and qPCR methodology has been described in details recently.[27 (link)28 ] Blood samples were irradiated, subjected to postirradiation incubation as described in section on “Sample preparation, radiation dose, and downstream processing” and total RNA was isolated directly from the blood using Trizol BD reagent (Sigma-Aldrich, USA) as per the manufacturer's recommendations. RNA sample (1 µg) was converted into cDNA using the high-capacity DNA-archive kit (Applied Biosystems, USA) and following the manufacturer's instructions. qPCR was done using the gene-specific TaqMan™ assays (Applied Biosystems, USA). The gene expression change has been calculated as fold change utilizing the “ΔΔCt” method. Thus, the gene expression fold change is expressed as 2−ΔΔCt. The data were evaluated using the Sequence Detection Software 1.3.1 (Applied Biosystems, USA). Statistical evaluation of the generated data was performed using one-way ANOVA for both the irradiated samples with respect to controls as well as the individual samples with respect to each other. The 18S-rRNA and GAPDH TaqMan™ assays were selected as the endogenous controls or normalizers.[27 (link)]
Nongrum S., Vaiphei S.T., Keppen J., Ksoo M., Kashyap E, & Sharan R.N. (2017). Identification and Preliminary Validation of Radiation Response Protein(s) in Human Blood for a High-throughput Molecular Biodosimetry Technology for the Future. Genome Integrity, 8, 5.
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2

Quantitative RT-PCR Gene Expression Analysis

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Total RNAs were extracted using TRI Reagent (Sigma). Samples were treated with DNase-1 (Ambion) and retro-transcribed using the High Capacity DNA Archive Kit (Applied Biosystems). PCRs were carried out using the SYBR Green PCR Master Mix on an ABI 7500HT machine (Applied Biosystems). Relative quantification was done using the ddCT (Pfaffl) method. Primer sequences are available upon request.
Forte G., Grossi V., Celestini V., Lucisano G., Scardapane M., Varvara D., Patruno M., Bagnulo R., Loconte D., Giunti L., Petracca A., Giglio S., Genuardi M., Pellegrini F., Resta N, & Simone C. (2014). Characterization of the rs2802292 SNP identifies FOXO3A as a modifier locus predicting cancer risk in patients with PJS and PHTS hamartomatous polyposis syndromes. BMC Cancer, 14, 661.
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3

Total RNA Extraction and cDNA Synthesis

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Total RNA was isolated by Qiazol reagent (Qiagen) following the manufacturer’s instructions. To avoid possible DNA contaminations, RNA was treated with DNase I (Ambion). RNA purity was checked by spectrophotometer and RNA integrity by examination on agarose gel electrophoresis. cDNA was synthesized retrotranscribing 4 μg of total RNA in a total volume of 100 μL using a High Capacity DNA Archive Kit (Applied Biosystems) for the manufacturer’s instructions.
Cariello M., Peres C., Zerlotin R., Porru E., Sabbà C., Roda A, & Moschetta A. (2017). Long-term Administration of Nuclear Bile Acid Receptor FXR Agonist Prevents Spontaneous Hepatocarcinogenesis in Abcb4−/− Mice. Scientific Reports, 7, 11203.
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4

RNA Extraction and cDNA Synthesis

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Total RNA was extracted from the PBMCs pellet by QIAzol® Lysis Reagent (Qiagen, Hilden, Germany), according to the manufacturer’s instructions using standard and previously validated protocol [11 (link)]. To avoid possible DNA contamination, the RNA was treated with DNAase-1 (Ambion, Foster City, CA). RNA purity (A260/A280 > 1.75), and the concentration was checked by spectrophotometer, while the RNA integrity was assessed by Bio-RAd ExperionTM (Bio-Rad, Hercules, CA). Only the samples with Relative Quality Index (RQI) > 8 were used for reverse-transcription. According to the manufacturer’s instructions, cDNA was synthesized by reverse-transcribing 4 μg of total RNA in a volume of 100 μl using the High Capacity DNA Archive Kit (Applied Biosystems, Foster Cyti, CA). For TaqMan gene validations, due to the small quantity of RNA achieved, we used a High Capacity RNA-to-cDNA Kit (Applied Biosystems, Foster City, CA) to reverse-transcribe 10 ng of total RNA in a volume of 20 μl.
Sangineto M., Graziano G., D’Amore S., Salvia R., Palasciano G., Sabbà C., Vacca M, & Cariello M. (2018). Identification of peculiar gene expression profile in peripheral blood mononuclear cells (PBMC) of celiac patients on gluten free diet. PLoS ONE, 13(5), e0197915.
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5

Quantifying Muscle Gene Expression Changes

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Approximately 50 mg of tissue from gastrocnemius muscle was cut on dry ice. Muscle RNA was isolated using qiazol reagent (Qiagen, Germantown, MD, USA) and reverse transcribed to cDNA (High Capacity DNA Archive Kit, Applied Biosystems; Thermo Fisher Scientific, Inc., Waltham, MA, USA). The cDNA was amplified by real-time quantitative PCR using pre-designed and validated primers under universal cycling conditions defined by Applied Biosystems (Thermo Fisher Scientific, Inc., Waltham, MA, USA). The primers used were (4331182 Murf-1 Mm01185221_m1; Atrogin1 Mm00499523_m1; Bax Mm00432051_m1; Bcl-2 Mm00477631_m1; Bnip3 Mm01275600; Ppargc1a Mm01208835; Ppargc1b Mm00504730; Nrf1 Mm01135606 ). The thermocycling conditions were 95°C for 10 min, followed by 40 cycles at 95°C for 15 sec, and 60°C for 1 min. Target gene expression was normalized to the endogenous control GAPDH (Product no.4352932E, Thermo Fisher Scientific, Inc., Waltham, MA, USA) in the same reaction and expressed as 2−ddct relative to the CON(−) group [36 (link)].
Snoke D.B., Nishikawa Y., Cole R.M., Ni A., Angelotti A., Vodovotz Y, & Belury M.A. (2021). Dietary naringenin preserves insulin sensitivity and grip strength and attenuates inflammation but accelerates weight loss in a mouse model of cancer cachexia. Molecular nutrition & food research, 65(22), e2100268.
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6

Total RNA Extraction and miRNA Isolation

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Total RNA was extracted by Qiazol reagent (Qiagen) following the manufacturer’s instructions. cDNA was synthesized retrotranscribing 4 μg of total RNA in a total volume of 100 μL using a High Capacity DNA Archive Kit (Thermo Fisher Scientific) for the manufacturer’s instructions. MiRNA isolation was performed using mirVana™ miRNA Isolation Kit (Thermo Fischer, MA, USA) following the manufacturer’s guidelines. Plasma miRNA isolation was performed using miRNaesy serum/plasma advanced kit (Qiagen, GmbH) following the manufacturer’s guidelines. RNA quality was assessed by 260 nm absorbance with Nanodrop spectrophotometer (NanoDrop Technologies Inc., Wilmington, DE, USA) and by capillary electrophoresis with an Agilent2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). For miRNA expression analysis, reverse transcription was made using TaqMan microRNA Reverse Transcription Kit (Thermo Fisher Scientific, MA, USA), following the manufacturer’s instructions.
Cariello M., Piccinin E., Pasculli E., Arconzo M., Zerlotin R., D’Amore S., Mastropasqua F., Peres C., Graziano G., Villani G., Pesole G, & Moschetta A. (2022). Platelets from patients with visceral obesity promote colon cancer growth. Communications Biology, 5, 553.
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7

Total RNA Isolation and cDNA Synthesis

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Total RNA was isolated by a Qiazol reagent (Qiagen) following the manufacturer’s instructions. To avoid possible DNA contaminations, the RNA was treated with DNase I (Thermo Fisher Scientific). RNA purity was checked by spectrophotometer and RNA integrity by examination on agarose gel electrophoresis. cDNA was synthesized retrotranscribing 4 μg of total RNA in a total volume of 100 μL using a High Capacity DNA Archive Kit (Thermo Fisher Scientific) following the manufacturer’s instructions.
Cariello M., Piccinin E., Zerlotin R., Piglionica M., Peres C., Divella C., Signorile A., Villani G., Ingravallo G., Sabbà C, & Moschetta A. (2021). Adhesion of Platelets to Colon Cancer Cells Is Necessary to Promote Tumor Development in Xenograft, Genetic and Inflammation Models. Cancers, 13(16), 4243.
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8

RNA Isolation and Reverse Transcription

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Total RNA was isolated by QIAzol Lysis Reagent (Qiagen) following manufacturer's instructions. To avoid possible DNA contamination, RNA was treated with DNAase-1 (Ambion, Foster City, CA). RNA purity was checked by spectrophotometer, while RNA integrity was assessed by Biorad Experion. Only samples with Relative Quality Index (RQI)>8 were used for reverse-transcription. According to the manufacturer's instructions, cDNA was synthesized by reverse-transcribing 4 µg of total RNA using the High Capacity DNA Archive Kit (Applied Biosystem).
Vacca M., D'Amore S., Graziano G., D'Orazio A., Cariello M., Massafra V., Salvatore L., Martelli N., Murzilli S., Sasso G.L., Mariani-Costantini R, & Moschetta A. (2014). Clustering Nuclear Receptors in Liver Regeneration Identifies Candidate Modulators of Hepatocyte Proliferation and Hepatocarcinoma. PLoS ONE, 9(8), e104449.
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9

Quantifying Gene Expression in Mice

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RNA was isolated from liver and ileum of mice using RNeasy Micro kit (Qiagen, Milano, Italy). cDNA was generated from 4 μg total RNA using High Capacity DNA Archive Kit (Applied Biosystem, Foster City, CA, USA) and following the manufacturer’s instructions. Primers to detect the mRNA expression level of each gene were designed using Primer Express software (Applied Biosystem) based on Gene Bank sequence data. mRNA expression levels were quantified by qRT-PCR using Power Sybr Green chemistry and normalized to cyclophilin mRNA levels for the DSS experiment and Gapdh for the AOM-DSS experiments and ApcMin/+ mice. Validated primers for qRT-PCR are available upon request. Real time qPCR dataare calculatedusing the Ct of basal condition or control as calibrator. Then, we represent relative quantification as mean of replicates.
Cariello M., Zerlotin R., Pasculli E., Piccinin E., Peres C., Porru E., Roda A., Gadaleta R.M, & Moschetta A. (2022). Intestinal FXR Activation via Transgenic Chimera or Chemical Agonism Prevents Colitis-Associated and Genetically-Induced Colon Cancer. Cancers, 14(13), 3081.
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10

RNA Extraction and qRT-PCR Analysis

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Total RNA was extracted using TRI reagent (Sigma) according to the manufacturer’s instructions. Samples were retro-transcribed using the High Capacity DNA Archive Kit (Applied Biosystem). PCRs were carried out in triplicate using the SYBR Green PCR Master Mix (Bio-Rad) on a CFX96 Touch Real-Time PCR Detection System (Bio-Rad), according to the manufacturer’s instructions. Relative quantification was done using the ddCT method. Primer sequences are available on request.
Peserico A., Germani A., Sanese P., Barbosa A.J., Di Virgilio V., Fittipaldi R., Fabini E., Bertucci C., Varchi G., Moyer M.P., Caretti G., Del Rio A, & Simone C. (2015). A SMYD3 Small-Molecule Inhibitor Impairing Cancer Cell Growth. Journal of cellular physiology, 230(10), 2447-2460.
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