Typer version 4

Totally, 4 variants (rs1048943, rs4646422, rs762551 and rs2470890) were selected in the global population on the basis of the 1000 Genomes Project (https://www.internationalgenome.org/) [17 (link)]. The minor allele frequency of every SNP was greater than 5%. Primers design for amplification and extension of SNPs listed in Supplementary Table 1 were completed by the Agena Bioscience Assay Design Suite V2.0 software (https://agenacx.com/online-tools/). Genomic DNA was extracted from the participants’ blood samples using the GoldMag-Mini Whole Blood Genomic DNA Purification Kit (GoldMag. Co. Ltd., Xi’an, China) regarded as the amplification template. Target amplification of SNPs was performed according to a predetermined PCR procedure, and PCR product purification was completed by agarose gel electrophoresis. After that, SNP genotyping was performed by the Agena MassARRAY platform with iPLEX gold chemistry (Agena Bioscience, San Diego, CA, USA), and data management was completed by Agena Bioscience TYPER, Version 4.0 [18 (link)].
Fig. 1

Linkage disequilibrium (LD) analysis of five SNPs in CYP1A1, and CYP1A2. The LD value is determined by r² > 0.8 analyzed by Haploview software, version 4.2

In our study, six SNPs (rs75267932, rs7336610, rs72640334, rs17735387, rs7318578, and rs1428) of the MIR17HG gene were selected by 1000 Genomes Project with MAF > 5% and r² (the measure value of linkage disequilibrium (LD)) < 0.8 and for further genotyping. The genomic DNA from each peripheral blood sample was extracted by a whole-blood genomic DNA extraction kit (GoldMag, Xi'an, China). The NanoDrop 2000C spectrophotometer (Thermo Scientific, Waltham, USA) was performed to test the concentration and purity of the genomic DNA. PCR primers used for genotyping were designed by the Agena Bioscience Assay Design Suite software (V2.0, https://agenacx.com/online-tools/). We further identified the SNP genotyping via the Agena MassARRAY iPLEX version 4.0 platform, and the data was organized and analyzed by the Agena Bioscience TYPER version 4.0 software.

Genomic DNA was extracted from whole blood samples with GoldMag whole blood genomic DNA purification kit (GoldMag Co. Ltd. Xi'an city, China). DNA concentration was evaluated with a NanoDrop 2000 platform (Thermo Fisher S 95% CIentific, Waltham, MA, USA), according to the manufacturer's instruction. The genes associated with TB were selected from the Genomic Wide Association Study using UCSC (http://genome.ucsc.edu/) database. We found that FOXO3 was associated with several diseases including TB (Greer & Brunet, 2008). The candidate SNPs of FOXO3 were selected from the 1,000 Genomes Project data (http://www.internationalgenome.org/) with minor allele frequency (MAF) > 0.02 in the Asian population and MAF > 0.05 in the Chinese Han Beijing population. Finally, four SNPs in FOXO3 were selected in this case–control study. The Agena Bioscience Assay Design Suite V2.0 software (https://agenacx.com/online–tools/) was performed to analyze the amplification and extension primers. The MassARRAY Nanodispenser and MassARRAY iPLEX platform (both from Agena Bios 95% CIence, San Diego, CA, USA) were used to genotype the SNPs. Data management was performed using Agena Bioscience TYPER version 4.0 software as previously described (Zhu et al., 2017).

In this study, five SNPs (rs3093203, rs3093144, rs12459936, rs3093110, and rs3093193) in CYP4F2 were selected according to previously published studies on the association between CYP4F2 polymorphisms and disease susceptibility (12 (link)–14 (link)). The genotype distributions of the candidate SNPs in controls met Hardy–Weinberg equilibrium (HWE) (p >0.05). All the candidate SNPs had a minor allele frequency (MAF) of >5% in the Han Chinese in Beijing (CHB) population from the 1,000 Genomes Project (http://www.internationalgenome.org/). The primers for five SNPs were designed by Agena Bioscience Assay Design Suite version 2.0 software. The polymorphisms were genotyped using the Agena MassARRAY platform (Agena Bioscience, San Diego, CA, USA) with iPLEX gold chemistry. Ultimately, Agena Bioscience TYPER version 4.0 software was used for data management and genotyping result analysis.