Request for quotation for next-generation sequencing (NGS) service in 2026

Provide comprehensive services for Next-Generation Sequencing (NGS) gene sequencing, as part of the procurement plan: Implement the project: "Application of Liquid Chromatography-Mass Spectrometry (LC-MS/MS) and Next-Generation Sequencing (NGS) techniques in the study of drug pharmacokinetics and DPYD genotype polymorphism related to the toxicity of fluoropyrimidine drugs in cancer patients," funded by the National Science and Technology Development Fund.

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All in one

1. **Introduction:** NGS-based genotyping: Complete package. The entire technical process, instruction manuals, experimental results, quality control samples, and related documents will be fully transferred to Party A upon contract completion. Party B is responsible for supporting implementation, training, and operating the procedure at Party A's facility. 2. **Technical Transfer:** Party B will perform the following transfers: * **2.1 Technical Procedure:** * DNA extraction protocol suitable for the analysis target. * DNA quantification and quality assessment protocol. * Library preparation protocol (if applicable). * Target gene capture protocol (if applicable). * Instrument operation protocol. * Quality control (QC) protocols at each stage of the process. * Data analysis from raw data to variant results. * Data storage and test result management. * Forms, tracking records, and implementation guides. * **2.2 Documentation:** * Detailed technical instruction manual (SOP). * Data analysis and interpretation guidelines. * Training materials and related quality control forms. * Illustrative experimental data and sample results report (if available). 3. **Training and Practical Guidance:** * Party B will provide training for 02 trainees designated by Party A. * The training content includes: * Principles and applications of next-generation sequencing technology for target genes. * Full process from DNA extraction to result delivery. * Operation of equipment and handling common situations. * Data analysis and interpretation of variants. * Quality control steps during implementation. * Training format: Direct training combined with practical exercises on real samples. * After completing the training, trainees will be able to independently perform the procedure under the supervision of the unit. 4. **Acceptance Criteria:** * Complete transfer of technical procedures and related documents. * Completion of training for 02 trainees according to the agreed-upon content. * Trainees can perform the main steps of the process and analyze sample data under the guidance of the transferring expert. * Experimental results meet the quality criteria specified in the transferred procedure. 5. **Technical Specifications:** * 1. DNA extraction and fragmenting from blood samples: * Total DNA is obtained using the GeneJET Genomic DNA Purification Kit (Thermo Scientific). * The DNA extraction kit uses a silica membrane column in a centrifuge format. * Suitable for various types of blood, including fresh or refrigerated animal blood (treated with EDTA, Citrate, etc.), bone marrow, and body fluids. * The process takes only 20 minutes after cell lysis and yields purified DNA with the required concentration and purity. * The extracted DNA is ready for use in all applications such as PCR, real-time PCR, Sanger sequencing, next-generation sequencing, etc. * 2. DNA fragmenting, end repair, and A-tailing: * DNA fragments are generated using the IGT® Enzyme Plus Library Prep Kit V3. * Uses Fragment & ERA Buffer v3 and an enzyme mixture containing Fragment & ERA Enzyme mix. * The enzyme mixture contains endonuclease that cuts double-stranded DNA into fragments with a size of 200–400 bp, while also having enzymes to perform end repair and A-tailing. * Suitable for DNA extracted from various types of samples. * 3. Adapter ligation and library amplification: * IGT® Adapter & Universal 10nt Primer 1–384 is a set of adapters and primers used in NGS library construction, which is used after adapter attachment in the library preparation process. This kit uses the Universal 10-nt Index to identify samples during multiple target gene sequencing. * Allows running multiple samples simultaneously while ensuring accurate sample identification. * Compatible with various types of libraries: whole genome, exome, targeted genes, etc. * 4. Target gene enrichment using TargetSeq® DNA Probes HT, TargetSeq One® Hyb & Wash Kit V2.0 and TargetSeq® Cap Beads: * TargetSeq® DNA Probes HT: A collection of biotinylated DNA probes specifically designed for 3 target genes (DPYD, TYMS, and ENOSF1) to be surveyed. The probe is used in the hybridization step to pair with library fragments containing target sequences. * Probes specific to gene regions (surveying >600 SNPs on 3 genes). * Stored at -20°C. * TargetSeq One® Hyb & Wash Kit V2.0: A chemical solution for performing the following steps: * Hybridization of DNA library and probe * Capture of target genes that have been hybridized * Washing away non-specific binding * PCR amplification of target genes * Stored at -20°C. * TargetSeq® Cap Beads: Streptavidin-coated beads used to retrieve DNA library fragments bound to biotin, increasing the accuracy of target gene capture and purification. * Streptavidin-coated beads * Stored at 2–8°C. * 5. Quantification using Qubit Fluorometer: * Qubit Fluorometer is a nucleic acid quantification device based on fluorescence principles, which can accurately quantify DNA or library concentration. * Qubit 1X dsDNA HS (High Sensitivity) Assay Kit is a DNA double-strand quantification reagent. * Fluorescent dye specifically binds to DNA strands. * The intensity of the fluorescence is proportional to the amount of DNA in the sample. * 6. Sequencing on MiSeq: * Library is modified with NaOH for MiSeq. * For MiSeq, the library must be converted into single-stranded DNA before being attached to the flow cell. * Single-stranded DNA is attached to oligonucleotides on the flow cell and amplified into clusters. **Key Parameters:** * Cluster density: >80% (recommended) * Q30: 75–85% (recommended) * 7. Data analysis using CLC Genomics Workbench (QIAGEN, Germany): * The sequencing data obtained from the MiSeq system is processed and analyzed using the CLC Genomics Workbench software (QIAGEN, Germany). Raw reads are quality-controlled, adapter sequences and low-quality bases are removed before alignment to the human reference genome (hg19). Variants such as single nucleotide variants (SNVs) and insertions/deletions (InDels) are detected using the Variant Detection module of the software. Variants are annotated based on public databases such as ClinVar, dbSNP, and gnomAD, and then filtered and analyzed according to research objectives.

Can Tho University of Medicine and Pharmacy