Theoretically, Illumina NGS technology resembles Sanger sequencing. During the cyclic process of DNA synthesis sequencing, a DNA polymerase catalyzes the incorporation of fluorescently labeled deoxyribonucleotide triphosphates (dNTPs) onto a DNA template strand. Each cycle, as the dNTP is incorporated, the nucleotide is identified via fluorescence excitation. However, diverging from Sanger sequencing, which sequences the entire completed DNA segment, NGS has revolutionized the process through its high-throughput parallel sequencing method.

•  Flow cell 

A core sequencing reactor vessel where DNA fragments attach to lanes on the surface. Each lane has adapters that match the adapters added to the ends of the DNA fragments. 

• Workflow 

The workflow includes library preparation, sequencing, analysis, and sharing. 

• Applications 

The NextSeq 550 System can be used for RNA sequencing, mRNA sequencing, gene expression profiling, and whole-exome sequencing.

The facility is made in such a way so that the entire work flow of the genomics study, that involves multiple steps viz, library preparation, quantitation and sequencing, through utilization of multiple instruments, can be performed within the facility itself.

Both the internal users and external users can avail the facility for usage. However the complete sequencing set-up is not yet chargeable because of the costly reagents required for preparing the libraries and the kits required for setting up the run in the NextSeq 550.

Right now, we are only charging the sequencing instrument on the data size generated. 

Major Equipment

• Sequencing Platform : Illumina NextSeq 550 from Illumina
• 4200 Tape Station system from Agilent
• Qubit 4 Fluorometer by Thermo Fisher Scientific
• Analysis workstation

Illumina Nextseq 550

The NextSeq 550 System combines tried-and-true next-generation sequencing (NGS) and array capabilities with tunable outputs, enabling both small and large labs to scale to meet their needs. Its fast DNA-to-results workflow enables rapid sequencing of exomes, targeted panels, and transcriptomes in a single run, with the flexibility to switch to low- or high-throughput sequencing as needed. Illumina scientists are available at every point along the way with support and guidance, enabling large clinical research labs to scale with confidence and smaller labs to employ both genotyping and sequencing technologies. 

The NextSeq 550 System offers various throughput levels, providing users with optimal operational efficiency. The NextSeq 550 System also delivers a one-day turnaround for numerous popular sequencing applications. With this instrument, users can sequence a broad range of samples per run: 

• 1-12 exomes

• 1-16 transcriptomes

• 6-96 targeted panels 

• 12-40 gene expression profiling samples

The NextSeq 550 System features an intuitive user interface and load-and-go design that allow users to perform various sequencing applications with minimal training and instrument set-up time. It provides Array Scanning

     The NextSeq 550 System offers:

    • Ready-to-use reagents 

    • Radio-frequency identification (RFID)–encoded consumables 

    • Integrated cluster generation and sequencing capabilities 

    • Easy-to-follow controls 

4200 Tape Station system from Agilent 

• Simplified workflow and minimal hands-on time with fully automated sample processing and the proven, ready-to-use ScreenTape technology
• Nucleic acid analysis with minimal manual intervention and excellent reproducibility for size, concentration, and integrity
• Wide range of RNA and DNA ScreenTape applications, including assays for genomic and cell-free DNA
• Easy switching between DNA and RNA ScreenTape assays for excellent application flexibility
• Full sample scalability from 1 to 96 samples with constant cost-per-sample
• Minimized time to results with as little as 1 to 2 minutes per sample or less than 90 minutes for 96 samples
• Reduced sample consumption with as little as 1 to 2 µL of DNA and RNA samples, even for high-sensitivity assays
• No risk of cross contamination with individual ScreenTape lanes
• Reliable integrity standards for RNA (RNA integrity number equivalent, RINe), genomic DNA (DNA integrity number, DIN), and cell-free DNA (%cfDNA)
• Optimized cost of ownership and maximized instrument uptime with the option for self-maintenance tasks

Focused-ultrasonicator_Covaris

• The M220 Focused-ultrasonicator enables precise and accurate results with AFA-energetics. Acoustic basedfocused shearing is performed at ultrasonic range of 500Khz.  Thus it functions beyond audible range ensuring no discomfort to operators.
• The M220 Focused-ultrasonicator system is able to process 1 sample per run in a minimal time.
• The M220 Focused-ultrasonicator has the capacity to handle a dynamic sample volume range for different sample types i.e. from 15 μl to 1 ml.
• The M220 Focused-ultrasonicator complies with the following key features such as random fragmentation (no fragmentation bias), dynamic range of fragment size (100bp - 5Kbp), isothermal process (no heat-induced bias or sample degradation), concentration independent, uniform genome coverage, high sample reproducibility and sample recovery during the whole process.
•  M220 system can generate 150bp DNA fragments within processing times as low as 5 minutes.

Applications:

a. Mechanical shearing of DNA for Next Generation Sequencing. 

b. DNA/RNA extraction from Formalin-Fixed, Paraffin-Embedded (FFPE) tissue samples. 

c. Extract DNA for NGS from whole blood. 

d. Chromatin Mechanical Shearing for ChIP-Seq.