Saliva samples from each lung transplant recipient were collected using the OGR-500 collection kit (DNA Genotek, Ottawa, ON, Canada). The median time to DNA collection was 144 days (range 9–1417 days) after transplantation. Genotyping was accomplished using a TaqMan® single tube genotyping assay using allelic specific primers (Life Technologies, Foster City, CA) for indicated SNPs. Allelic PCR results were analyzed using Taqman Genotyper v1.0.1 (Life Technologies) reported by DNA Genotek and entered into the REDCap database at Washington University.
Taqman genotyper v1
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Taqman Genotyper v1.0.1 is a software application designed for the analysis of genetic data. It provides tools for visualizing and interpreting genotyping data generated from Taqman-based assays.
Automatically generated - may contain errors
Lab products found in correlation
Quantstudio 12k flex instrument, by Thermo Fisher Scientific (3 mentions)
Allelic specific primers, by Thermo Fisher Scientific (2 mentions)
Ogr 500 collection kit, by DNA Genotek (2 mentions)
Gentra puregene blood kit, by Qiagen (1 mentions)
Biophotometer, by Eppendorf (1 mentions)
Geneamp pcr system 9700, by Thermo Fisher Scientific (1 mentions)
Abi prism 7900ht sequence detection system, by Thermo Fisher Scientific (1 mentions)
Taqman open array genotyping system, by Thermo Fisher Scientific (1 mentions)
Oragene dna saliva collection kit, by DNA Genotek (1 mentions)
Infinite m200 pro multimode reader, by Tecan (1 mentions)
Taqman real time pcr assay, by Thermo Fisher Scientific (1 mentions)
8 protocols using taqman genotyper v1
1
Genotyping of Lung Transplant Recipients
2
Genotyping of Lung Transplant Recipients
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Saliva samples from each lung transplant recipient were collected using the OGR-500 collection kit (DNA Genotek, Ottawa, ON, Canada). The median time to DNA collection was 144 days (range 9–1417 days) after transplantation. Genotyping was accomplished using a TaqMan® single tube genotyping assay using allelic specific primers (Life Technologies, Foster City, CA) for indicated SNPs. Allelic PCR results were analyzed using Taqman Genotyper v1.0.1 (Life Technologies) reported by DNA Genotek and entered into the REDCap database at Washington University.
3
Genetic Profiling of Dementia Risk
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Genomic DNA was extracted from peripheral blood, and whole-genome amplified, fragmented, hybridized, fluorescently tagged, and scanned using the Infinium assay [21 (link)]. Sixty-four SNPs were selected based on previous associations with dementia, cognition, neuroanatomical differences, and blood pressure (Supplementary Table 1 ), and further selected in our sample. Genomic DNA was normalized to a concentration of ~ 50 ng/μl, and 2.5 μL of genomic DNA was mixed with 2.5 μL TaqMan OpenArray Master Mix. The resulting samples were dispensed using the OpenArray® AccuFill™ System onto OpenArray plates with each plate containing 48 samples and 65 SNP assays per sample. The QuantStudio™ 12K Flex instrument (Applied Biosystems, Carlsbad, CA, USA) was used to perform the real-time PCR reactions on the loaded OpenArray plates. The fluorescence emission results were read using the OpenArray® SNP Genotyping Analysis software v1 (Applied Biosystems), and the genotyping analysis performed using TaqMan® Genotyper v1.3 with the auto call feature and the default settings.
4
Genotyping SNPs in Livestock Genes
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Genomic DNA was extracted using the Gentra Puregene blood kit (Qiagen, Hilden, Germany), and purity and concentration were measured with an Eppendorf BioPhotometer (Eppendorf, Hamburg, Germany). A custom open array, based on the TaqMan real-time PCR assay, was designed for single nucleotide polymorphism (SNP) genotyping. It included a total of 8 SNPs: 2 from the SPP1 gene, 3 from POFUT1 and 3 from PRLR (Table 1 ). Context sequences are given in Supplementary Information Table S1 . Genotyping was carried out using a 12 K Flex QuantStudio instrument (Thermo Fisher Scientific, Waltham, MA, USA). Genotypes were visualized with Taqman Genotyper v.1.3 software (Applied Biosystems, Waltham, MA, USA).
The Haploview software package [34 (link)] was used to estimate and plot pairwise linkage disequilibrium (LD) measures (D′ and r2) and to infer haplotype frequencies as well as to define LD blocks according to the Gabriel criteria [35 (link)]. Haploview was also used to estimate minor allele frequencies (MAF) and observed and expected heterozygosites and to identify significant departures from the Hardy–Weinberg equilibrium at each polymorphic locus.
The Haploview software package [34 (link)] was used to estimate and plot pairwise linkage disequilibrium (LD) measures (D′ and r2) and to infer haplotype frequencies as well as to define LD blocks according to the Gabriel criteria [35 (link)]. Haploview was also used to estimate minor allele frequencies (MAF) and observed and expected heterozygosites and to identify significant departures from the Hardy–Weinberg equilibrium at each polymorphic locus.
5
Genetic Variant Genotyping Protocol
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File Builder software (Thermo Fisher Scientific) was used to design the forward (5′-GGCCAGCAGGAGGACAAG-3′) and reverse (5′-GTTGCCCTCGCCAGTGA-3′) primers as well as VIC (5′-AGGACACAGACGTCCACT-3′) and FAM (5′-AGGACACAGACTTCCACT-3′) labeled probes. The assay was validated on control samples obtained from the proband and his family, and then the carrier frequency study was performed on 1,021 samples. The samples were run on the GeneAmp® PCR System 9700 (Thermo Fisher Scientific) at the following PCR conditions: hold at 50°C for 2 minutes and 95°C for 10 minutes, then 40 cycles at 95°C for 15 seconds and 60°C for 1 minute. Allelic discrimination was then performed on the ABI PRISM® 7900 HT Sequence Detection System (Thermo Fisher Scientific), and data were analyzed using TaqMan Genotyper v1.2 software (Thermo Fisher Scientific).
6
DNA Genotyping Protocol for Genetic Studies
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A biological sample was obtained from each participant using the Oragene DNA saliva collection kit (OG-500; DNA Genotek Inc, Ottawa, ON, Canada). DNA extraction was performed by using standard procedures. DNA concentration was measured by absorbance using the Infinite® M200 PRO multimode reader (Tecan US Inc, Research Triangle Park, NC, USA).
Genotyping was performed using TaqMan® OpenArray™ Genotyping System (Thermo Fisher Scientific, Waltham, MA, USA) following the manufacturer’s instructions. Raw data were analyzed with the TaqMan® Genotyper v1.2 software (Thermo Fisher Scientific).
Stringent quality control criteria were applied to both SNP and individual data. SNPs were excluded if they had a missing genotype rate higher than 1% or showed departure from the Hardy–Weinberg equilibrium (P<0.01). Individuals with genotypic data showing a missing rate >10% or those with a non-European ancestry were also excluded.
After quality control procedures, 85 SNPs and 567 individuals were finally included in the analyses. Detailed information about those SNPs, such as location, function, or possible allelic variants, is given inTable S1 .
Genotyping was performed using TaqMan® OpenArray™ Genotyping System (Thermo Fisher Scientific, Waltham, MA, USA) following the manufacturer’s instructions. Raw data were analyzed with the TaqMan® Genotyper v1.2 software (Thermo Fisher Scientific).
Stringent quality control criteria were applied to both SNP and individual data. SNPs were excluded if they had a missing genotype rate higher than 1% or showed departure from the Hardy–Weinberg equilibrium (P<0.01). Individuals with genotypic data showing a missing rate >10% or those with a non-European ancestry were also excluded.
After quality control procedures, 85 SNPs and 567 individuals were finally included in the analyses. Detailed information about those SNPs, such as location, function, or possible allelic variants, is given in
7
Genotyping Goat Y-Chromosome SNPs
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Seven single nucleotide polymorphisms (SNPs) located in the non-recombining region of the goat Y-chromosome were genotyped according to [16 (link)], i.e., SRY-2971 T > A, SRY-3098 G > A, SRY-1876 A > C (at the sex determining region Y gene), AMELY-42 C > T (amelogenin, Y-Linked, GenBank AY082491) and ZFY-527 A > G (zinc finger protein, Y-Linked, GenBank AY082500). These markers have been reported in previous studies [14 (link),15 (link),28 ,29 (link)], while markers DDX3Y g.56 C > G (DEAD-box helicase 3, Y-linked) and ZFY g.46 C > T were described for the first time by Vidal et al. [16 (link)]. A custom TaqMan Real-Time PCR assay was designed and genotypic data were generated on a 12K Flex QuantStudio instrument (Thermo Fisher Scientific) at the Servei Veterinari de Genetica Molecular from the Universitat Autònoma de Barcelona [30 ]. Genotypes were visualized with the Taqman Genotyper v.1.3 software (Applied Biosystems) and SNPs with call rates < 0.9 were removed from the dataset.
8
Sheep Genome SNP Genotyping
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A custom TaqMan Real-Time PCR assay (Thermo Fisher Scientific, Waltham, MA) was designed for SNP genotyping. It included a total of 31 SNP mapping to the sheep GHR gene. The GHR SNP were retrieved from the Ensembl Genome database (http: / / www .ensembl .org/ Ovis _aries/ Gene/ Variation _Gene/ Table ?db = core;g = ENSOARG00000008837;r = 16: 31829583 -32003795;t = ENSOART00000009622). Moreover, 2 and 3 SNP mapping to the GHRHR and IGF1, respectively, were retrieved from the dbSNP database (https: / / www .ncbi .nlm .nih .gov/ projects/ SNP/ ) and genotyped in the 380 Sarda sheep. Sequences flanking the SNP are given in Supplemental Table S1 (https: / / doi .org/ 10 .3168/ jds .2018 -14914) and the full panel of 36 SNP is reported in Table 1. Each SNP plus 60-bp flanking sequences were submitted to the Custom TaqMan Assay Design Tool web page (https: / / www .thermofisher .com/ order/ custom -genomic -products/ tools/ genotyping/ ) to design an assay matching the producer's criteria. Genotyping was carried out in the Servei Veterinari de Genetica Molecular from the Universitat Autonoma de Barcelona (http: / / sct .uab .cat/ svgm/ en) by using a 12K Flex QuantStudio instrument (Thermo Fisher Scientific). Genotypes were visualized with the Taqman Genotyper v.1.3 software (Applied Biosystems, Foster City, CA), and SNP with call rates <0.9 were removed from the data set.
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