Services

DNA Sequencing is a High-throughput approach to determining a DNA molecule's exact sequence of bases. DNA Sequencing is extensively used to identify disease-causing variants/ changes, infer the genomic sequence of an organism, and genotyping.

*Low input options are available. Input requirement is the available standard option.

^$sequencing depth recommendation is for the equivalent human genome.

Any RNA isolation method or kit is acceptable to use provided that the final isolate is total RNA, intact, and free of contaminants, including DNA, excess salts, proteins, and RNAses. Total RNA is always run on the YCGA bioanlayzer prior to starting the library prep. When a sample is found to have low purity or integrity the user will be notified and asked if they would like either to continue or resubmit. Please note that YCGA is not responsible for poor data resulting from samples that did not pass initial qc but were approved for processing by the user.

Experiments should be performed with two or more biological replicates (preferably 3 replicates), unless there is a compelling reason why this is impractical or wasteful. A biological replicate is defined as an independent growth of cells/tissue and subsequent analysis. Technical replicates from the same RNA library are not required, except to evaluate cases where biological variability is abnormally high.

The first option to consider when submitting your samples is the library prep method. All of our library prep services start with total RNA and generate ready to sequence cDNA libraries. YCGA offers the following RNA library prep services:

1. Ribosomal reduction: This process efficiently removes the rRNA component of total RNA using a hybridization/bead capture procedure that selectively binds target sequences using biotinylated capture probes. The process minimizes ribosomal contamination and optimizes the percentage of reads covering RNA species. This library prep is ideal for partially degraded RNA or for sequencing non-polyadenylated transcripts. > 200ng of total RNA should be submitted.
2. PolyA selection: This process specifically selects/enriches for polyadenylated transcripts using oligo-dT beads followed by random priming. This process is ideal for samples that are not degraded (RIN > 7) and for sequencing applications that only require data from polyadenylated transcripts. > 200ng of total RNA should be submitted.
3. Low-input poly A selection: Provides an efficient solution for generating high-quality cDNA libraries for RNA-seq from very low amounts of input RNA. This library prep is not strand-specific and is only recommended when less than 50ng of a sample is available.
4. FFPE RNA: This library prep converts total RNA into template molecules of known strand origin, followed by sequence-specific capture of coding RNA. This provides a low-cost solution for analyzing human RNA isolated from FFPE (formalin-fixed, paraffin-embedded) tissues and other low-quality samples. RNA requires a DV200 value of 30% or higher. 20-100ng of RNA is required depending on DV200.
5. Micro RNA: This library prep is ideally suited to isolate small RNA transcripts and convert them into barcoded cDNA libraries ready for sequencing. > 200ng of total RNA should be submitted.
6. Custom/User-Provided RNA sequencings such as RIP-Seq and Ribosome profiling. Please inquire with the facility.
7. Digital droplet PCR: Instrument is suitable for absolute quantification of gene expression, copy number and allele frequency in a multiplex format. Up to 5 independent assays can be tested in a single well.

^$sequencing depth recommendation is for the equivalent human genome. For denovo transcript assembly and isoform identification we recommend 80-100 million reads/ sample

1. 1ATAC-Seq: to study accessible regions/open chromatin regions in the chromosome.
2. HiC: to construct a three-dimensional organization map of chromosome
3. DNA Methylation: Methylation sequencing interrogates the epigenetic information mediated by cytosine modifications (5mC and 5hmC) through enzymatic or bisulfite conversion of unmethylated Cytosine followed by sequencing in the NGS platform. The following services are available for methylation studies.
   1. Whole genome methylation: sequencing of the entire genome to identify the methylation status of Cytosine.
   2. Methyl-Seq: Targeted Capture and Sequencing to identify methylation status of Cytosine in DMRs, GENCODE promoters, CpG islands, shores, shelves, and DNase hypersensitive sites.
   3. Custom Targeted Methylation: Targeted Capture and Sequencing to identify the methylation status of Cytosine in the genomic region based on specific experimental questions.

*Low input options are available. Input requirement is available standard option.

^$sequencing depth recommendation is for human equivalent genome.

Cell-free DNA extraction, whole genome library preparation, Exome capture, methylation status, and fragmentomics are offered as services. We prepare these libraries from low input material (as low as 6ng total input).

We offer Sequencing services using MiSeq, Novaseq 6000, NextSeq2000, Novaseq xPlus, and Element AVITI. Our standard service offering is paired-end 150 bp (300 cycles). Investigators are free to request a specific number of reads per their requirements. We also sequence user-prepped libraries with unique sequencing requirements.

We offer multiple spatial technologies for expression analysis and epigenetic profiling. Contact Bony De Kumar to learn more about spatial and single-cell technologies and choose an appropriate technology for your needs.