Abi 3730xl dna analyzer platform

We selected representative virus strains and variants for complete genome sequencing. Specific primers for amplification and sequencing are listed in Additional file 1 [27 (link)], and primer synthesis was performed by a qualified third-party biotech company. The PCR product was purified using a QIAquick gel extraction kit (Qiagen) according to the manufacturer’s instructions and then sent to a qualified third-party biotech company for sequencing using an ABI 3730xl DNA analyzer platform (Applied Biosystems, CA, USA). Nucleotide sequences were initially assembled with Lasergene software (version 8.0; DNASTAR Inc., Madison, WI, USA), and continuous sequences were aligned using BioEdit software (version 7.0.5). In addition, selected strains were transferred to a third-party biotech company (BGI, China) for complete genome sequencing by next-generation sequencing.

The ERG11 gene encoding lanosterol 14α-demethylase was amplified and sequenced with specific primers (Table S1). The BigDye^® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems, USA) was used for sequencing the reaction, precipitated with ethanol/EDTA/sodium acetate according to the protocol suggested by the manufacturer and sequenced on the ABI3730xl DNA Analyzer platform (Applied Biosystems, USA). DNA sequences and the corresponding amino acid sequences were analyzed with the SeqMan II and Bioedit software packages (Lasergene; DNAStar, USA). Consensus sequences were aligned with different reference ERG11 sequences from Candida spp. For the mutation analysis, the C. albicans strain SC5314 ERG11 gene assembly sequence from Candida Genome Database (http://www.candidagenome.org/) was used [31 (link),32 (link),33 (link)].

Genomic DNA was extracted with a Gentra^® Puregene^® Yeast and G+ Bacteria Kit (Qiagen^®, Hilden, Germany). Hot spot (HS) regions, HS1 and HS2, were amplified with the following forward and reverse primers (1 μM concentration of each): HS1-F, 5′-TGTTCCTTTCGCTTACGCCT-3′; HS2-F, 5′-GGGTGAGCAGATGTTGTCCA-3′; HS1-R, 5′-ACGACCGATGGAGAACACAC-3′; HS2-R, 5′-CCCATGTAGATGGCGGAGTC-3′. BigDye^® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems, Waltham, MA, USA) was used for the sequencing reaction, precipitated with ethanol/EDTA/sodium acetate according to the protocol suggested by the manufacturer and sequenced on an ABI3730xl DNA Analyzer platform (Applied Biosystems, Waltham, MA, USA). DNA sequences and the corresponding amino acid sequences were analyzed with the SeqMan II and Bioedit 7.2 software packages (Lasergene—DNAStar, Madison, MI, USA). The sequence of the FKS1 (orf 19.2929) gene from the Candida Genome Database (CGD; http://www.candidagenome.org; accessed on 10 January 2023) and from C. haemulonii B11899 were used as a reference [20 (link),21 (link)].