Yeastar genomic dna isolation kit

For genomic DNA isolation, a glycerol stock of Wyeast 3068 strain received from the brewers was streaked onto rich media (YPD) agar plates and incubated for 3–5 days. A single colony was isolated and cultured for DNA isolation. Genomic DNA extraction was performed using Zymo YeaStar Genomic DNA Isolation kit as per manufacturer recommendations. Genomic DNA was sheared to an average length of 410 bp before sequencing on an Illumina NextSeq 500 platform. Sequences were first analyzed using FastQC to assess overall quality. The 1.6 million read pairs were then mapped to the Saccharomyces cerevisiae S288C reference genome R64 (sacCer3) using bowtie2 (v 2.2.6) in local alignment mode. We observed a high overall mapping rate of 92% with low rates of duplication (0.5%) and reads mapping to multiple locations (4.2%). A moderate rate of indel detection (13.4%) and the relatively low 68.8% of reads mapped as proper pairs suggested this commercial yeast differs from the lab strain in some structural ways. Sequencing reads are deposited in the NCBI Sequence Read Archive (SRA) under BioProject PRJNA1011390.

All the molecular biology enzymes, P. pastoris codon optimized EDIII gene of dengue virus serotype-1 (DENV-1) with 6×-His tag at 3′end, zeocin antibiotic, LIVE/DEAD FungaLight yeast viability kit, 96-DWP (square wells with V-shaped bottom; 2.2 ml total volume), breathable rayon tape, anti-His mAb and UltraPure DNase/RNase-free distilled water were purchased from Thermo Fisher Scientific Corporation, USA. 96-DWP (square wells with U-shaped bottom; 2.2 ml total volume per well) was from Genetix Biotech Asia Pvt Ltd, India. Goat anti-mouse IgG-H&L-chain was procured from Jackson ImmunoResearch Laboratories, Inc. USA. N1-europium chelate was synthesized at University of Turku, Finland. YeaStar genomic DNA isolation kit was purchased from Zymo Research, CA, USA. 2x SSO EvoGreen mix, hard-shell white 96-well PCR plate with clear wells and microseal ‘B’ adhesive sealing film were obtained from Bio-Rad Laboratories, CA, USA. All the other chemicals were procured from Sigma-Aldrich Corporation, USA and culture media and casamino acids were purchased from Becton, Dickinson and Company, USA. The casamino acids (CA) is acid hydrolysed casein with low sodium chloride and iron concentrations (Bacto^TM casamino acids Cat # 223050). Peptone is an enzymatic digest of animal protein (Bacto^TM peptone Cat # 211677). Primers for quantitative PCR (qPCR), were synthesized by IDT, Singapore.

For genomic DNA isolation, a glycerol stock of Wyeast 3068 strain received from the brewers was streaked onto rich media (YPD) agar plates and incubated for 3–5 days. A single colony was isolated and cultured for DNA isolation. Genomic DNA extraction was performed using Zymo YeaStar Genomic DNA Isolation kit as per manufacturer recommendations. Genomic DNA was sheared to an average length of 410 bp before sequencing on an Illumina NextSeq 500 platform. Sequences were first analyzed using FastQC to assess overall quality. The 1.6 million read pairs were then mapped to the Saccharomyces cerevisiae S288C reference genome R64 (sacCer3) using bowtie2 (v 2.2.6) in local alignment mode. We observed a high overall mapping rate of 92% with low rates of duplication (0.5%) and reads mapping to multiple locations (4.2%). A moderate rate of indel detection (13.4%) and the relatively low 68.8% of reads mapped as proper pairs suggested this commercial yeast differs from the lab strain in some structural ways. Sequencing reads are deposited in the NCBI Sequence Read Archive (SRA) under BioProject PRJNA1011390.