Fragment Analysis (FA), also known as Fragment Length analysis (FLA), is a powerful technique used in a wide-range of applications—detection of mutations, genotyping, identifying short tandem repeats, and DNA profiling. Eurofins Genomics offers highest accuracy in sequencing the fully prepared samples via capillary electrophoresis using ABI's 3730xl DNA analyzers. Our current offering in Fragment Analysis includes the Ready2Load service. 

Order

Ready2Load Service

The Fragment Analysis - Ready2Load service allows you to get your assay samples analyzed on our ABI DNA analyzers.

What you do

• Design your fragment analysis assay using the dyes from DS-33 (see below)
• Perform PCR based on your assay design
• Assemble your sample(s) (dye-labeled PCR Product + DNA Size-standards + Hi-Di Formamide)
• Ship the assembled samples to Eurofins Genomics in a secure 96-well microtiter plate (see below)

What we do

• Load and run the samples on ABI 3730xl DNA analyzer
• Send the sequence data to your email

Please note that we currently samples from assays using the following dye combination.

Additional Information

When all that is remaining to complete your Fragment Analysis assay is for the samples to be run on a ABI 3730xl DNA Analyzer, ship your sample to Eurofins Genomics! The Ready2Load Fragment Analysis service offered by Eurofins Genomics includes the following services:

• Accurate separation of fragments by Capillary Electrophoresis (CE) on ABI 3730xl sequencing machine under standard conditions
• Sequence data sent via email as .fsa files (use GeneMapper or GenoTyper analysis software to read the same).
• Typical turn around time for the delivery of .fsa files is within 3 days.
• Analysis of fragments spanning longer lengths will be considered on request

Dye-Labeled DNA fragments can be analyzed on the Applied Biosystems 3730xl DNA analyzers. PCR amplification of a Dye-labeled primer is the most widely used method for incorporating dyes in DNA fragments. 

Several genetic analysis experiments incorporate the dye in DNA fragments to gather critical information about the organisms. The applications of fragment analysis that we can process include include SNP Genotyping, Amplified Fragment Length Polymorphism (AFLP), loss of heterozygosity and INDEL evaluations. 

Please note that we currently only accept samples labeled with the dyes in the DS-33 Matrix Standard Kit (Dye Set G5), i.e., 6-FAM, VIC, NED, PET and LIZ. The 600 LIZ® DNA Size Standard must be added to you Ready2Load samples. Please adhere to the guidelines in the Sample Submission Guidelines document.

Sample Submission Guidelines for Fragment Analysis

Eurofins Genomics (US) offers the ‘Fragment Analysis – Ready2Load’ service using ABI 3730xl DNA analyzers. Our workflow is optimized to offer the service for the PCR products generated by microsatellite, SNP, LOH, AFLP, RFLP, T-RFLP, ARISA or other protocols.

Dye Set

Only the following dye combinations can be used as matrix and for labelling the PCR products:
ABI Standard Dye Set DS-336-FAM, VIC, NED, PET and LIZ*
*the dye LIZ is typically reserved for the size standard and should not be used for PCR product labelling.Size Standard

The protocol for the Ready2Load fragment analysis at Eurofins Genomics US is optimized for analysis of DNA fragments 50–600 bp. The LIZ 600 size standard must be included in your samples for fragment analysis.

Sample preparation

Please adhere to the following guidelines, to assemble samples for Fragment Analysis – Ready2Load service.

• Run PCR using dye labeled primers that can be analyzed with ABI Standard DS-33 filter set
• Prepare solutions of PCR products at 50–100 ng/μL
• Assemble the samples for ‘Fragment Analysis – Ready2Load’ Service, by mixing the following:
  1. Hi-Di Formamide loading buffer, 10 μL (Applied Biosystems)
  2. Dye-Labeled internal size standard, 0.5 μL (use only LIZ-600).
  3. PCR product, 2 μL
• The total sample volume should be 10–15 μL.

Plate Type

• 96-well plates (MicroAmp® Optical 96-Well Reaction Plate, ThermoFisher, catalogue no. 4316813, or equivalent)
• Alternatively, customers may submit in 1.75 mL centrifuge tubes. Orders are still submitted online and must meet the minimum number of samples to qualify. Pricing is the same for either method.

Concentration

• Concentration across all samples submitted in an order must be normalized
• For multiplex analysis, the concentrations of the PCR products in a sample must be normalized in order to get a balanced profile (recommended variations in concentration <50%)

Shipping

Ship the liquid samples at ambient temperature to:

ATTN: Fragment Analysis Sequencing Lab
Eurofins Genomics
12701 Plantside Drive,
Louisville, KY 40299

• Use 8-cap strips for sealing 96-well plates (recommended)
• Seal the plate tightly and avoid folding; don’t freeze and thaw the samples after sealing.

How to Order

To place an order for ‘Fragment Analysis – Ready2Load’ service, visit the order page.

1. Go to the order page
2. Download the excel upload template
3. Add sample names to the upload sheet.
4. Upload the template
5. Click next
6. Add to cart

A note about sample concentration

• Concentration across all samples submitted in an order must be normalized
• For multiplex analysis, the concentrations of the PCR products in a sample must be normalized in order to get a balanced profile (recommended variations in concentration <50%)

Contact

Please do not hesitate to contact us in case of any questions! Email: GenomicsSupport@eurofins.com Phone: 1-800-688-2248