Quant it picogreen dsdna assay kit

Manufactured by Qiagen

Sourced in United States

The Quant-iT™ PicoGreen™ dsDNA Assay Kit is a fluorescence-based reagent used for the quantification of double-stranded DNA (dsDNA) in solution. The kit provides a sensitive and accurate method to measure dsDNA concentrations.

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Lab products found in correlation

Allprep dna rna protein mini kit, by Qiagen (3 mentions) Epitect bisulfite kit, by Qiagen (2 mentions) Biosprint 96 dna plant kit, by Qiagen (2 mentions) Rnaeasy mini kit, by Qiagen (1 mentions) Agilent 2100 bio analyzer agilent rna 6000 nano kit, by Agilent Technologies (1 mentions) Rnalater, by Qiagen (1 mentions) Qiaamp dna mini kit, by Qiagen (1 mentions) Qiamp fast dna stool mini kit, by Qiagen (1 mentions) Miseq system, by Illumina (1 mentions)

8 protocols using quant it picogreen dsdna assay kit

Methylation Profiling of Cord Blood

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We collected whole blood samples of cord blood within 30 min after delivery by research staff, as described previously [21 (link)]. Samples were then aliquoted (300–500 µL/aliquot) and stored at − 80 °C. Double strand DNA concentration was assessed using Quant-iT™ PicoGreen™ dsDNA Assay Kit (Qiagen, USA) after DNA extraction using the All Prep DNA/RNA/Protein Mini Kit (Qiagen, USA). Methylation levels at > 850,000 CpG loci were quantified using the Infinium MethylationEPIC BeadChip after sodium-bisulfite conversion [21 (link)]. Preprocessing was preformed using the minfi R package [26 (link)]. We removed non-CpG probes, probes that were annotated to sex chromosomes, probes that showed non-significant detection (p value > 0.05) in ≥ 5% of the samples, probes with a single nucleotide polymorphism (SNP) with a minor allele frequency ≥ 5% at the target CpG or at the single base extension, as well as cross-reactive probes reported in [27 (link)]. We used the ComBat function in the sva R package to account for batch effects [28 (link)]. We retained a total of 790,563 high quality probes for statistical analyses.

Lu T., Cardenas A., Perron P., Hivert M.F., Bouchard L, & Greenwood C.M. (2021). Detecting cord blood cell type-specific epigenetic associations with gestational diabetes mellitus and early childhood growth. Clinical Epigenetics, 13, 131.

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DNA and RNA Isolation Protocols

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DNA was isolated from peripheral whole blood lymphocytes using the DNA extraction 500 arrow® Kit (DiaSorin Ireland Ltd) kit. DNA was quantified with NanoDrop (NanoDrop Products Thermo Scientific Wilmington, DE) and by fluorometric reading (Quant-iT™ PicoGreen® dsDNA Assay Kit).
RNA was extracted from PBMCs (Mononuclear cell fractions, isolated over a Ficoll-Hypaque density gradient) using Qiagen RNAeasy mini kit (Qiagen, Hilden, Germany) and quantified using NanoPhotometer (NanoPhotometer™Pearl, Denville®, Denville Scientific, Holliston, MA).

Loi E., Moi L., Fadda A., Satta G., Zucca M., Sanna S., Amini Nia S., Cabras G., Padoan M., Magnani C., Miligi L., Piro S., Gentilini D., Ennas M.G., Southey M.C., Giles G.G., Wong Doo N., Cocco P, & Zavattari P. (2019). Methylation alteration of SHANK1 as a predictive, diagnostic and prognostic biomarker for chronic lymphocytic leukemia. Oncotarget, 10(48), 4987-5002.

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Placental DNA and RNA Extraction

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Placental samples were collected after delivery (<30 min) by well-trained research staff. A total of 1 cm³ was collected on the fetal side, near the umbilical cord insertion (≤5 cm) and Gen3G samples were kept in RNAlater (Qiagen, Germantown, MD, USA). Samples from both cohorts were kept at −80 °C until DNA extraction. DNA and RNA were extracted using the All Prep DNA/RNA/Protein Mini Kit (Qiagen, Germantown, MD, USA) for both cohorts. Double strand DNA concentration were assessed using Quant-iT™ PicoGreen™ dsDNA Assay Kit (Qiagen, Germantown, MD, USA). Sodium-bisulfite conversions were performed prior to epigenome-wide methylation analyses following recommendations of the EpiTect Bisulfite Kit (Qiagen, Germantown, MD, USA). RNA concentrations and RNA integrity number (RIN) (mean 8.3 ± 0.9) were assessed with Agilent 2100 Bioanalyzer (Agilent RNA 6000 Nano Kit, Agilent Technologies, Santa Clara, CA, USA). Samples with RIN < 6 were excluded.

Gagné-Ouellet V., Breton E., Thibeault K., Fortin C.A., Desgagné V., Girard Tremblay É., Cardenas A., Guérin R., Perron P., Hivert M.F, & Bouchard L. (2020). Placental Epigenome-Wide Association Study Identified Loci Associated with Childhood Adiposity at 3 Years of Age. International Journal of Molecular Sciences, 21(19), 7201.

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Optimized MDA Protocol for HCV Genome

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In order to facilitate experimental optimization, we defined a standard MDA protocol with a reaction volume of 40 μL and consisting of phi29 DNA polymerase and its corresponding buffer, 1 mM of dNTPs, and 2 μM or 50 μM of random exonuclease-resistant primers. A recombinant 12-kb plasmid (HCV plasmid) containing a 9-kb HCV genotype 1a genome from our previous study (20 (link)) was used as a positive control. The reaction was incubated at 30°C for 16 h, followed by heat-inactivation at 65°C for 10 min. In addition to visualization on a 0.8% agarose gel, MDA yield was quantitated by the Quant-iT PicoGreen dsDNA Assay kit (QIAGEN, Valencia, CA) after purification using the QIAamp DNA mini kit. Due to the extreme sensitivity of the techniques (5 (link)), all MDA experiments were performed in strict adherence to the recommendations of Champlot et al. to avoid possible contamination (21 (link)).

Wang W., Ren Y., Lu Y., Xu Y., Crosby S.D., Di Bisceglie A.M, & Fan X. (2017). Template-dependent multiple displacement amplification for profiling human circulating RNA. BioTechniques, 63(1), 21-27.

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Gut Microbiota Profiling in Mice

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The gut microbiota from OP and OR mice fed on chow or high-fat diet for 24 h were measured by sequencing 16S rRNA from feces. Fecal gDNA was extracted from 100 mg of feces per animal using a QIAmp Fast DNA Stool Mini Kit (Qiagen) according to the manufacturer’s instructions and quantified using the Quant-iT Picogreen dsDNA Assay Kit. The 16S rRNA V4-V5 regions were amplified using the Platinum Taq DNA Polymerase High Fidelity enzyme. For the PCR reaction, 100 ng of gDNA was used in conjunction with 10 μM of primers V4F (5’-TCG GCA GCG TGC MGC CGC GGT AA-3’) and V5R (5’-GTCTCG TGG GCT CTT YMT TTR AGT TT-3’), in a total reaction volume of 25 μL. The PCR cycling program was 30 s at 94 °C; 30 cycles of 15 s at 94 °C, 30 s at 62 °C, 45 s at 68 °C; hold at 4 °C. The product was purified with AMPURE XP reagent and quantified with the Quant-iT Picogreen dsDNA Assay Kit. The samples were prepared for 16S ribosomal RNA gene amplicons for the Illumina MiSeq System, and 4 nmol of the samples were used for Miseq Illumina analyses using the V2 2x250bp kit according to the manufacturer’s instructions.

Razolli D.S., de Araújo T.M., Sant’Ana M.R., Kirwan P., Cintra D.E., Merkle F.T, & Velloso L.A. (2019). POMC processing in the hypothalamus is directly regulated by saturated fat − implications for the development of obesity. Neuroendocrinology, 110(1-2), 92-104.

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Sorghum Genomic DNA Extraction and Accession Curation

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The HBP (N = 296) are inbred lines derived from a recurrent selection breeding population developed by intercrossing germplasm that survived natural SCA infestation. For genomic DNA extraction, fresh leaf tissue of each accession was collected from 2-week-old seedlings raised in a greenhouse. Tissue was lyophilized for 2 days and then ground using a 96-well plate plant tissue grinder (Retsch Mixer Mill). Genomic DNA was extracted using the BioSprint 96 DNA Plant Kit (QIAGEN), quantified using the Quant-iT PicoGreen dsDNA Assay Kit, and normalized to 10 ng/μl. An additional set of global accessions (GDP, N = 767) was assembled on the basis of published datasets (57 (link), 58 (link)), selecting the subset of accessions with georeference information and other passport data such as country of origin and morphological type (fig. S1 and file S1). The GDP accessions were from 52 countries on five continents and all major botanical types including 164 caudatum, 96 guinea, 81 durra, 57 bicolor, and 47 kafir accessions; 288 of other botanical types; and 34 accessions of unknown botanical type. A subset of publicly available pre-breeding lines (N = 35) from Texas A&M University’s sorghum program was identified from the GDP.

Muleta K.T., Felderhoff T., Winans N., Walstead R., Charles J.R., Armstrong J.S., Mamidi S., Plott C., Vogel J.P., Lemaux P.G., Mockler T.C., Grimwood J., Schmutz J., Pressoir G, & Morris G.P. (2022). The recent evolutionary rescue of a staple crop depended on over half a century of global germplasm exchange. Science Advances, 8(6), eabj4633.

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Placental DNA/RNA Extraction and Bisulfite Conversion

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We extracted DNA and RNA from placental biopsies using the All Prep DNA/RNA/Protein Mini Kit (Qiagen, USA) following the manufacturer’s standard procedure and quantified double-strand DNA using Quant-iT™ PicoGreen™ dsDNA Assay Kit (Qiagen, USA). We preformed sodium-bisulfite conversion of DNA prior to the methylation analyses following recommendations of the EpiTect Bisulfite Kit (Qiagen, USA).

Gagné-Ouellet V., Breton E., Thibeault K., Fortin C.A., Cardenas A., Guérin R., Perron P., Hivert M.F, & Bouchard L. (2020). Mediation Analysis Supports a Causal Relationship between Maternal Hyperglycemia and Placental DNA Methylation Variations at the Leptin Gene Locus and Cord Blood Leptin Levels. International Journal of Molecular Sciences, 21(1), 329.

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Genomic DNA Extraction from Sorghum Accessions

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The HBP (N = 296) are inbred lines derived from a recurrent selection breeding population developed by intercrossing germplasm that survived natural SCA infestation. For genomic DNA extraction, fresh leaf tissue of each accession was collected from two weeks old seedlings raised in a greenhouse. Tissue was lyophilized for two days and then grounded up using a 96-well plate plant tissue grinder (Retsch Mixer Mill). Genomic DNA was extracted using the BioSprint 96 DNA Plant Kit (QIAGEN), quantified using Quant-iT™ PicoGreen® dsDNA Assay Kit, and normalized to 10 ng/uL. An additional set of global accessions (GDP, N = 767) was assembled based on a published data set (50, 51) including sorghum accessions from 52 countries on five continents and all major botanical types (Supp. Fig. S1, Supp. File S1). The GDP accessions included 164 caudatum, 96 guinea, 81 durra, 57 bicolor, and 47 kafir accessions, along with 288 of other botanical types and 34 accessions of unknown botanical type.

Muleta K.T., Felderhoff T., Winans N.D., Walstead R., Charles J.R., Armstrong J.S., Mamidi S., Plott C., Vogel J.P., Lemaux P.G., Mockler T.C., Grimwood J., Schmutz J., Pressoir G., & Morris G.P. (2021). The recent evolutionary rescue of a staple crop depended on over half a century of global germplasm exchange.

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