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Massarray analyzer

Manufactured by Agena
Sourced in United States

The MassARRAY analyzer is a high-throughput, mass spectrometry-based platform designed for genetic analysis. It is capable of accurately detecting and quantifying DNA or RNA targets in a multiplex format.

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10 protocols using massarray analyzer

1

DNA Methylation Analysis of ADAMTS9

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Genomic DNA of BGC-823 and AGS cells were extracted using QIAamp DNA Mini Kit (Qiagen). Then, the methylation of ADAMTS9 promoter was detected using an MassARRAY analyzer (Agena Bioscience, San Diego, CA, USA). EpiTYPER software was used to calculate the methylation ratio.
Other experimental and statistical methods are available in the Supplementary dataset.
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2

Quantifying CpG Methylation in p16 and p21 Promoters

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To quantify the CpG methylation status within p16Ink4a and p21Cip1/Waf1 promoters in MEFs, bisulfite conversion of genomic DNA was performed using an EZ DNA methylation kit (Zymo Research) as described previously [47 (link)]. PCR amplification of the targeted promoter regions was performed using the primers listed in Supplementary Table 1. After in vitro transcription using the PCR products as the template and T7 RNA polymerase, uracil-specific cleavage of the RNA transcripts by RNase A was conducted and the sizes of the resulting fragments were determined by MALDI-TOF mass spectrometry (MassARRAY Analyzer, Agena Bioscience, San Diego, CA, USA). The data were analyzed using an EpiTYPER software.
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3

Genotyping BDNF rs6265 Polymorphism

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Genomic DNA was extracted from the stored EDTA anticoagulated-venous blood using a DNA purification kit (Puregene, Gentra Systems, Minneapolis, MN, United States). Then, the genotype of BDNF rs6265 was detected using the Sequenom method (Beijing Cnkingbio Biotechnology Corporation, Beijing, China). The following forward and reverse primers were used: 5′-ACGTTGGATGTTGT TTTCTTCATTGGGCCG-3′ and 5′-ACGTTGGATGGCTT GACATCATTGGCTGAC-3′, respectively. The target region was amplified using PCR Accessory Set (Sequenom, 11327). The PCR products were processed using iPLEX Gold reagents and purified using a SpectroCHIP Resin kit (Sequenom, 10117-2). MassARRAY® Analyzer (Agena, 260000) was used to acquire data from the purified samples. No inconsistency was found when 10% of the samples were randomly selected and genotyped again.
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4

Genotyping by Mass Spectrometry

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DNA was extracted from blood samples using DNeasy Blood and Tissue kit (Qiagen). The genotyping was performed using a multiplexed primer extension (SBE) chemistry of the iPLEX assay with detection of the incorporated allele by mass spectrometry with a MassARRAY analyzer from Agena Bioscience. Raw data from the mass reader was converted to genotype data using the Typer software (Agena Bioscience). Reproducibility was controlled to be 100%. The tool PLINK (version 1.07)64 (link) was used for Chi-square calculations to test for association between SNV allele and phenotype. The experiment was performed twice.
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5

Genotyping of CA6 gene SNPs

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Single nucleotide polymorphisms in the region of the CA6 gene were selected and genotyped on DNA isolated from whole stimulated saliva. The criteria for SNP selection were a) minor allele frequency (MAF) ≥ 5% in the CEU population b) not in complete linkage disequilibrium (r2 ≤ 0.8) and c) in the 10,000 bp up-stream to 10,000 bp down-stream region of the CA6 gene. Using the NIH snptag resource (https://snpinfo.niehs.nih.gov/snpinfo/snptag.html) 30 SNPs met these criteria. Genotyping was performed using a multiplexed primer extension chemistry of the iPLEX assay with detection of the incorporated allele by mass spectrometry using a MassARRAY analyzer (Agena Bioscience, Hamburg, Germany). Raw data from the mass reader was converted to genotype data using the Typer software (Agena Bioscience)39 –41 (link). Two SNP markers had sample call rate 0%, rs2274333 and rs17032942, the reasons for the failure was low allele signals resulting in inaccurate genotype calls. An additional SNP (rs6697763) was excluded from analysis for deviation from Hardy-Weinberg equilibrium (p = 0.033), leaving 27 SNPs with valid genotype data, where the average call rate per sample was 99.4% and overall call rate was 99.8% (Table S1). Genotyping data are uploaded and available at figshare 10.6084/m9.figshare.6948020.
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6

Detailed FFPE DNA Extraction Protocol

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Each FFPE block was shaved to 75–100 µm thickness in a sterile microcentrifuge tube. The samples were then deparaffinized with xylene and absolute ethanol, respectively, before genomic DNA extraction using a DNeasy Blood and Tissue Kit (Qiagen, Hilden, Germany) following manufacturer’s instructions. The DNA concentration from each sample was quantified using a Nanodrop Lite Spectrophotometer (Thermo Scientific, MA, USA) and kept at -20 °C until use. The DNA samples were required to have a level of quality marked by at least a 260/280 absorbance ratio between 1.5 and 2.0 and concentration ≥ 5 ng/µl, when examined using a MassARRAY analyzer (Agena Bioscience, CA, USA).
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7

Genotyping using MassARRAY® Analyzer

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Following the addition of 41 µl of HPLC-grade water and conditioning with ion exchange resin, reaction mixtures were loaded into a SpectroCHIP Array and analysed using MassARRAY® Analyzer (Agena Bioscience, CA, USA).
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8

Genotyping of JLH Chicken SNPs

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Genomic DNA was extracted from Chinese-native Jin Ling Hua (JLH) chickens (n = 384), and its quality and quantity were determined using a NanoDrop spectrometer and agarose gel electrophoresis analysis. MassARRAY detection was performed by Beijing Compass Biotechnology Co., Ltd. (Beijing, China) using the MassARRAY® analyzer (Agena Bioscience, San Diego, CA) to characterize the genotypes of candidate SNPs. Briefly, specific amplification of candidate SNPs was performed in a Veriti® 384-Well Thermal Cycler (Applied Biosystems, Foster City, CA) and the PCR products treated with alkaline phosphatase. A single-base extension reaction and resin purification were carried out, after which the PCR products were hybridized to 384 chips for mass spectrometry detection. Finally, single SNP correlation analysis of 21 SNPs in a total of 384 individuals was performed using a general linear model with PLINK.
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9

Genotyping Panel for Personalized Oncology

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Panel for genotyping was designed to include 62 polymorphisms that presented potential implication in patients’ treatment efficacy or drugs toxicity based on the bibliographic search performed (design named VIP-Onco). We could obtain suitable samples from 64 patients, and their genotyping was carried out at Centro Nacional de Genotipado (CEGEN, Santiago de Compostela, Spain) by mass spectrometry, employing a MassArray Analyzer (Agena Bioscience, San Diego, CA, USA).
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10

Genotyping HFE Mutations via MassARRAY

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DNA samples were transferred on PCR plates and sent to the SNP&SEQ Technology Platform, Uppsala University (National Genomics Infrastructure [NGI], SciLifeLab Sweden). The genotyping was performed using a multiplexed primer extension (SBE) chemistry of the iPLEX assay with detection of the incorporated allele by mass spectrometry with a MassARRAY analyzer from Agena Bioscience.28 , 29 (link), 30 (link) Raw data from the mass reader were converted to genotype data using the Typer software (Agena Bioscience). Both HFE C282Y (rs1800562) and H63D (rs1799945) genotype distributions were in Hardy‐Weinberg equilibrium (Ps > .1).
Given the limited number of individual carriers of H63D or C282Y, we pooled the participants in 2 groups: non‐carriers of any of the mutations (HFE‐neg) and carriers of either the H63D and/or C282Y (HFE‐pos).
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