Purelink viral rna dna kit

Manufactured by Thermo Fisher Scientific

Sourced in United States

The PureLink Viral RNA/DNA Kit is a rapid and efficient solution for the simultaneous extraction and purification of viral RNA and DNA from a variety of sample types. The kit utilizes a silica-based membrane technology to capture and concentrate viral nucleic acids, allowing for reliable and consistent results.

Automatically generated - may contain errors

Lab products found in correlation

Close spelling variants of this product

PureLink RNA kit (191 citations) PureLink kit (67 citations) PureLink® Viral RNA/DNA (2 citations)

28 protocols using purelink viral rna dna kit

Quantifying SARS-CoV-2 on Mask Samples

Check if the same lab product or an alternative is used in the 5 most similar protocols

At the end of each experimental run, the target-containing bag was immediately transferred to the virology laboratory in the next room. The wells of each target were washed with 500 μL of phosphate-buffered saline; the solution was collected in sterile Eppendorf vials and stored at –80 °C until analysis. Samples with the same surface area as the target’s wells (1.9 cm²) were cut from the external layer of the mask, inserted in Eppendorf vials containing 500 μL of phosphate-buffered saline, and stored as previously described. The presence of HCoV-229E on targets was determined via quantitative real-time PCR. RNA extraction was performed with a PureLink viral RNA/DNA kit (ThermoFisher). A total of 500 μL of viral suspension was used, and the elution was performed with 10 μL of elution buffer. RNA was retrotranscribed with a SuperScript VILO cDNA synthesis kit (ThermoFisher) and amplified with an HCoV-229E–specific real-time PCR gene assay (assay ID, Vi06439671_s1; 4331182; ThermoFisher).

Ionescu A.C., Brambilla E., Manzoli L., Orsini G., Gentili V, & Rizzo R. (2021). Efficacy of personal protective equipment against coronavirus transmission via dental handpieces. Journal of the American Dental Association (1939), 152(8), 631-640.

+ Open protocol

+ Expand

Fecal Microbiome DNA Extraction and Sequencing

Check if the same lab product or an alternative is used in the 5 most similar protocols

DNA from fecal samples, at day 0 (B0) was extracted with E.Z.N.A.^® Stool DNA Kit (Omega Bio-Tech, Norcross, GA, USA). Two types of kit for the nucleic acid extractions from samples after 30 days (B1) of culture were used: MasterPure Complete DNA and RNA purification kit (Lucigen, WI, USA) and PureLink Viral RNA/DNA kit (Thermo Fisher Scientific, Waltham, MA, USA). All extractions were performed following the manufacturers’ recommendations. The Ovation^® Ultralow V2 DNA-Seq Library Preparation kit (NUGEN, San Carlos, CA, USA) was used for library preparation, following the manufacturer’s instructions. Both input and final libraries were quantified with the Qubit 2.0 fluorometer (Invitrogen, Carlsbad, CA, USA), and the quality was tested by the Agilent 2100 Bioanalyzer High Sensitivity DNA assay (Agilent Technologies, Santa Clara, CA, USA). Libraries were then prepared for sequencing and sequenced on NovaSeq 6000 in 150-bp paired-end mode. Bioinformatic reconstruction was calculated on the fasta file of bacterial sequences on UniProt. The results are presented in Table S1, in Supplementary Material.

Brogna C., Brogna B., Bisaccia D.R., Lauritano F., Marino G., Montano L., Cristoni S., Prisco M, & Piscopo M. (2022). Could SARS-CoV-2 Have Bacteriophage Behavior or Induce the Activity of Other Bacteriophages?. Vaccines, 10(5), 708.

+ Open protocol

+ Expand

Viral RNA/DNA Extraction and qPCR Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

The collected chromatographic fractions were initially treated with TurboDNAse followed by RNA isolation using a Purelink Viral RNA/DNA Kit (ThermoFisher Scientific, Waltham, MA). The samples were then combined with TaqMan fast virus, custom TaqMan probe, and the primers listed in Table 3, and analyzed using a CFX Duet Real-Time qPCR System (Bio Rad, Hercules, Ca).

Sundstedt A., Sigvardsson M., Leanderson T., Hedlund G., Kalland T, & Dohlsten M. (1996). In vivo anergized CD4+ T cells express perturbed AP-1 and NF-kappa B transcription factors.. Proceedings of the National Academy of Sciences of the United States of America, 93(3), 979-984.

+ Open protocol

+ Expand

SARS-CoV-2 Viral RNA Extraction from Plasma

Check if the same lab product or an alternative is used in the 5 most similar protocols

Plasma samples stored at −80°C were thawed at room temperature prior to extraction. Plasma volumes used for pelleting the virus were based on the sample viral loads. For samples with viral loads ≥4.5 log10 copies/ml, 500μl of plasma was used to pellet the virus. We used larger plasma volumes of 1000μl – 1500μl to pellet the virus for samples with viral loads <4.5 log10 copies/ml, to increase viral concentration and PCR success. Respective volumes for each sample were centrifuged at 23 000 × g for 1 hour at 4°C to pellet the virus.
The supernatant was discarded from each sample leaving 200μl of pelleted plasma sample for extraction. Viral RNA (vRNA) was extracted using a PureLink Viral RNA/DNA Kit (ThermoFisher Scientific, USA) according to manufacturer’s instructions. In summary, 25μl of proteinase K and 200μl of lysis buffer were added to 200μl of pelleted plasma sample, mixed by vortexing and incubated for 15 minutes at 56°C. Absolute ethanol (250μl) was added to the lysate, mixed by vortexing and incubated for 5 minutes at room temperature. The lysate was transferred to a viral spin column (Invitrogen, California, USA) and centrifuged at 6800 × g for 1 minute. The column was washed twice using 500μl of wash buffer and spun at 6800 × g for 1 minute discarding flow through. Viral RNA was eluted in 30μl of buffer and used immediately for reverse transcription.

Chimukangara B., Varyani B., Shamu T., Mutsvangwa J., Manasa J., White E., Chimbetete C., Luethy R, & Katzenstein D. (2016). HIV drug resistance testing among patients failing second line antiretroviral therapy. Comparison of in-house and commercial sequencing. Journal of virological methods, 243, 151-157.

+ Open protocol

+ Expand

Quantitative PCR Detection of HCoV-229E on Surfaces

Check if the same lab product or an alternative is used in the 5 most similar protocols

At the end of each experimental run, the target-containing bag was immediately transferred to the virological laboratory in the next room. The wells of each target were washed with 500 μl PBS; the solution was collected in sterile Eppendorf vials and stored at −80°C until analysis. Samples with the same surface area as the target’s wells (1.9 cm²) were cut from the mask’s external layer, inserted in Eppendorf vials containing 500 μl PBS, and stored as previously described. HCoV-229E presence on targets was assessed using Real-Time quantitative PCR (qPCR). RNA extraction was performed using the Purelink viral RNA/DNA kit (ThermoFisher, Milan, Italy). A total of 500 μl of viral suspension was used, and the elution was performed with 10 μl elution buffer. RNA was retrotranscribed with Superscript VILO cDNA synthesis kit (ThermoFisher) and amplified with an HCoV-229E specific qPCR gene assay (Vi06439671_s1, Catalog number: 4,331,182, ThermoFisher).

Ionescu A.C., Brambilla E., Manzoli L., Orsini G., Gentili V, & Rizzo R. (2021). Aerosols modification with H2O2 reduces airborne contamination by dental handpieces. Journal of Oral Microbiology, 13(1), 1881361.

+ Open protocol

+ Expand

Quantification of Viral RNA from Cells and Plasma

Check if the same lab product or an alternative is used in the 5 most similar protocols

RNA was purified from 10⁵ sorted cells kept in Trizol (Life Technologies) according to the manufacturer’s instructions. Viral RNA was extracted from 200 μl of plasma with PureLink Viral RNA/DNA Kit (Life Technologies). RNA was treated with Turbo-DNA free kit (Life Technologies) and quantified by RT-qPCR using 4X TaqMan Fast Virus 1-Step Master Mix (Life Technologies), 750 nM of primers and 200 nM of probe (S1 Table). 10-fold serial dilutions of a SIVmac251 plasmid including SIV gag gene were performed to generate a standard curve, starting at 10⁹ SIV genome copies/μl. Amplifications were carried out with a 7500 Real-Time PCR System (Applied Biosystems), using the following parameters: 50°C /5 min, 95°C /20 sec, 40 cycles (95°C /15 sec, 60°C /1 min). 18S rRNA was used as endogenous control using the Eukaryotic 18S rRNA Endogenous Control mix (Life Technologies) for cell samples. Results are expressed as SIV RNA copies per cell detected in cell samples or copies per μl in serum samples.

Moukambi F., Rabezanahary H., Rodrigues V., Racine G., Robitaille L., Krust B., Andreani G., Soundaramourty C., Silvestre R., Laforge M, & Estaquier J. (2015). Early Loss of Splenic Tfh Cells in SIV-Infected Rhesus Macaques. PLoS Pathogens, 11(12), e1005287.

+ Open protocol

+ Expand

Isolation of Cell-Free DNA Using PureLink Kit

Check if the same lab product or an alternative is used in the 5 most similar protocols

PureLink Viral RNA/DNA kit (Life Technologies) was used to isolate cell-free DNA (cfDNA) as per manufacturer’s instructions.

Gould M.P., Bosworth C.M., McMahon S., Grandhi S., Grimerg B.T, & LaFramboise T. (2015). PCR-Free Enrichment of Mitochondrial DNA from Human Blood and Cell Lines for High Quality Next-Generation DNA Sequencing. PLoS ONE, 10(10), e0139253.

+ Open protocol

+ Expand

Quantifying Influenza Virus Infection in Mice

Check if the same lab product or an alternative is used in the 5 most similar protocols

At 48 h after virus infection, mice were sacrificed and homogenates of the right lung lobes were prepared. Total RNA was isolated from the homogenates using TRIzol (Life Technologies, Carlsbad, CA, USA) according to the manufacturer's instructions and stored at −70°C. IAV nucleic acids in the cell culture supernatant were extracted using the PureLink Viral RNA/DNA kit (Life Technologies). Total RNA was reverse transcribed into cDNA using the Prime Script RT Master kit (Takara Bio) at 42°C for 15 min and 95°C for 5 min, and qRT-PCR was performed using the SYBR R Premix Ex Taq kit (Takara Bio, Otsu, Japan) using the following forward and reverse primers: H1N1, 5′-TTCTAACCGAGGTCGAAACG-3′ and 5′-ACAAAGCGTCTACGCTGCAG-3′; and β-actin, 5′-AAATCGTGCGTGACATCAAAG-3′ and 5′-AAGAAGGAAGGCTGGAAAAGAG-3′. The annealing temperature was 60°C for both reactions. All reactions were performed in triplicate. The mRNA expression level of IAV in the lung tissue of mice was normalized by subtracting the cycle threshold value of β-actin from that of IAV.

Shi L., Yin S., Ren H., Gao R., Ma Z., Sun W, & Cao J. (2022). Influenza A Virus Production in Mouse Lung Is Inhibited by Inhalation of Aerosol Polyethylenimine/Short Hairpin RNA Plasmid Complexes. Disease Markers, 2022, 7404813.

+ Open protocol

+ Expand

Quantifying SIV Viral Loads in RMs

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Viral loads in the sera of SIV-infected RMs were quantified by quantitative real-time PCR (RT-PCR) using a PureLink viral RNA/DNA kit (Invitrogen). Primers used were SIVmac-F, GCA GAG GAG GAA ATT ACC CAG TAC, and SIVmac-R, CAA TTT TA CCC AGG CAT TTA ATG TT. The probe used was SIVmac-Probe, 6FAM TGT CCA CCT GCC ATT AAG CCC GA TAMRA (6FAM, 6-carboxyfluorescein; TAMRA, 6-carboxytetramethylrhodamine). A plasmid encoding the gag gene of SIVmac251 was used as a standard. Amplifications were carried out with a 7500 real-time PCR system (Applied Biosystems) using the following parameters: 50°C for 5 min, 95°C for 20 s, and 40 cycles of 95°C for 15 s and 60°C for 1 min. Samples were run in duplicates, and results are expressed as SIV RNA copies/mL, with a limit of detection of 40 copies/mL (64 (link)).

Yero A., Farnos O., Clain J., Zghidi-Abouzid O., Rabezanahary H., Racine G., Estaquier J, & Jenabian M.A. (2022). Impact of Early ARV Initiation on Relative Proportions of Effector and Regulatory CD8 T Cell in Mesenteric Lymph Nodes and Peripheral Blood During Acute SIV Infection of Rhesus Macaques. Journal of Virology, 96(7), e00255-22.

+ Open protocol

+ Expand

Quantification of SIV/SHIV Viral Loads in RMs

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

As previously described (106 (link), 107 (link)), viral loads in the sera of SIV- and SHIV-infected RMs were quantified by RT-qPCR using a PureLink Viral RNA/DNA Kit (Invitrogen). The PCR mixture comprises 4× TaqMan Fast Virus 1-Step Master Mix (Applied Biosystems), 750 nM of primers and 200 nM of probe. Primers and probe sequences are listed in Table S3. Serial 10-fold dilutions of a plasmid encoding for gag gene of SIVmac251 were performed to generate a standard curve. Amplifications were performed with a QuantStudio 6 Flex Real-Time PCR System (Applied Biosystems), using the following parameters: 50°C/5 min, 95°C/20 s, and 40 cycles (95°C/15 s, 60°C/1 min). Samples were run in duplicate, and results expressed as SIV RNA copies per milliliter.

Clain J.A., Boutrais S., Dewatines J., Racine G., Rabezanahary H., Droit A., Zghidi-Abouzid O, & Estaquier J. (2023). Lipid metabolic reprogramming of hepatic CD4+ T cells during SIV infection. Microbiology Spectrum, 11(5), e01687-23.

+ Open protocol

+ Expand

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?

Revolutionizing how scientists
search and build protocols!