Sanger Sequencing Service

High accuracy, Precision analysis

The Core Facility provides Sanger DNA sequencing and fragment analysis services to researchers at  academic institutions, government agencies and private companies.Analyzers to provide customers with the most sophisticated capillary sequencing technology to date. We accept purified PCR products, plasmids and  DNA constructs.

Purified PCR/ Plasmid Sequencing

Sanger Based Sequencing

Process
•   Quality check(QC) of PCR product/Plasmid by gel electrophoresis for purity & integrity.
•   Sequencing of DNA with client specific primer or universal primer using Sanger sequencing chemistry.
Sample Requirement
Purified PCR product- 10-20μl of 50-100 ng/μl per sample,Plasmid- 25μl of 50-100ng/μl per sample with customer specified primers, conc.= 10 pmole/μl( μM), volume- 10 μl and Tm/Primer sequence details of the primers.
Turn Around Time
24 working Hours from the receipt of sample(s).
Deliverables
Q20 Read length up to 800 bases with ABI,PDF and FASTA file.
Technology
Analysis using ABI 3130xl DNA Analyzers for capillary electrophoresis and fluorescent dye terminator detection.

Unpurified PCR/ Plasmid Sequencing

Sanger Based Sequencing

Process
•   Quality check(QC) of Unpurified PCR DNA/Plasmid by gel electrophoresis for purity & integrity.
•   Sequencing of DNA with client specific primer or universal primer using Sanger sequencing chemistry.
Sample Requirement
Unpurified PCR product- 10-20μl of 50-100 ng/μl per sample
Unpurified Plasmid- 25μl of 50-100ng/μl per sample with customer specified primers, conc.= 10 pmole/μl( μM), volume- 10 μl and Tm/Primer sequence details of the primers.
Turn Around Time
36 working Hours from the receipt of sample(s).
Deliverables
Q20 Read length up to 800 bases with ABI,PDF and FASTA file.
Technology
Analysis using ABI 3130xl DNA Analyzers for capillary electrophoresis and fluorescent dye terminator detecti

r-Ecoli Culture Sequencing Service

Sanger Based Sequencing

Process:
• r-E. coli culture containing plasmid DNA, propagation in test tubes or in 96 deep well culture plates.
• Plasmid DNA isolation and its quality check.
• DNA sequencing with client specific primer or universal primer using Sanger sequencing Techniques.
Sample Requirement:
E.coli culture – 96-well bacterial culture in 200μl LB media /antibiotic markers /culture plates/ glycerol stock, details of inserted gene size and gene information, vector details, customer specified primers, conc.= 10 pmole/μl ( μM), volume- 10 μl and Tm/Primer sequence details of the primers.
Turn Around Time:
2-3 working days from the receipt of sample(s).
Deliverables:
Q20 read length up to 800 bases with ABI,PDF and FASTA file.

Primer Walk Sequencing Service

Sanger Based Sequencing

Process:
•   Primer design and synthesis.
•   DNA isolation and sequencing with primers.
•   Universal primer details used for sequencing.
•   Read length up to 800 bases with ABI,PDF and FASTA file.
•   Quality control analysis reports.
Sample Requirement:
4 to 8 μg of purified PCR product (1-4 kb) / plasmid (10 kb) with insert size up to 3-4 kb/glycerol stock of culture, details of inserted gene size and gene information, vector details, customer specified primers, conc.= 10 pmole/μl ( μM), volume- 20μl and Tm of the primers.
Turn Around Time:
Turn around time for primer walk will depend on number and size of the insert DNA.
Deliverables:
Read length up to 800 bases with ABI,PDF and FASTA file.

Microsatellite Genotyping Service

Sanger Based Sequencing

Process:  .
•  Isolation of DNA from your sample(s).
• Amplification of Microsatellite loci under study using fluorescently-labelled primer pairs.
•  High-throughput fragment analysis of labeled amplicon.
•  Data analysis and generation of reports.
Sample Requirement:
•  Purified labeled PCR product: 20μl of 10-20ng/μl per sample in 96-well plate.
•  DNA: 20μl of 150-200 ng/μl of gDNA .
Turn Around Time:
10-12 days from the receipt of sample(s).
Deliverables:
•  A report with a summary of the methods followed.
• The results of the bioinformatic analysis including a table with the alleles called at each locus in each individual.

SNP Genotyping by Sequencing

Sanger Based Sequencing

Sanger sequencing-based SNP genotyping can be performed on a small subset of samples or as part of large-scale projects.  Let the Sanger sequencing experts to handle your SNP genotyping research objectives with speed, quality, and accuracy.
Sample requirement:
2-5μg of gDNA with quality report ,primer information for amplification of locus in which SNP is to be identified with standardized conditions, SNP information per sample and reference sequence.
The process includes:
•   Assay primer design based on gene sequence from data base.
•   PCR amplification and purification of amplicon.
•   High-throughput automated DNA Sequencing.
•   Sequence trace alignment and editing.
Deliverables:
Raw data files consist of chromatogram, FASTA files and report containing SNP list with minor allelic frequency.

Microbial Identification

Sanger Based Sequencing

Microbial Identification Service is based on sequencing of the 16SrRNA gene (bacterial) and ITS/D2 region of 28S Large subunit (LSU) rRNA gene for (Fungal/Yeast/Algae) phylogenetic analysis.

Types of Microbial Identification Service:
MID-S: This service includes sequencing of ribosomal genes or other conserved region of up to 1500 bp and its sequencing data with chromatogram and FASTA file.

MID-C: This service includes sequencing of ribosomal genes or other conserved region of up to 1500 bp and its contig report with sequencing data consisting of chromatogram and FASTA file.

MID-R: This service includes sequencing of ribosomal genes or other conserved region of up to 1500 bp and its comprehensive report includes genus and species level identification with phylogenetic tree and distance matrix generated using MEGA 4.1 for representation of the relationships, to obtain organism lineage and taxonomy.

Process:

1) Genomic DNA extraction from microbial culture & amplification of the desired gene by PCR.
2) Enzymatic/Gel based purification of the amplicon.
3) Sequence data generation by primer walk using BDTv3.1 chemistry on ABI 3730xl.
4) Quality check, trimming of low quality sequence from ends and assembly.

Sample Requirement
a) Pure culture propagation: for Bacteria/Yeast/Fungi (Plate/Slants/Glycerol stock), culture growth conditions (medium / temperature)
b) Isolated microbial genomic DNA: Concentration of 100 – 200 ng/μl , volume:- 20 μl
c) PCR product of 16S/18S rDNA gene: concentration of 20-30 ng/μl, volume:- 20 μl.

Turn Around Time:
3-12 working days from the receipt of sample.

Deliverables:
The Report will include: Genus and species level identification with top ten matches and DNA consensus sequence along with electropherogram, phylogenetic tree and distance matrix generated using MEGA 4.1 for representation of the relationships, to obtain organism lineage and taxonomy.

 

Customized Services

Sanger Based Sequencing

Sanger sequencing services provide high-quality results, industry-leading customer service and fast turnaround times at competitive prices.
We are well-equipped to handle DNA sequencing of all complexities.
 Our customized service offers the following advantages:
• Advanced Technology
: Modified protocols based on template sequencing service; developed in-house.
• Individual Consultation: Specialized consultation and customized solutions for your specific DNA sequencing needs
• Technical Support