Fragment Analysis Service
Fragment analysis run is an ABI3730xl electrophoresis run of 96 samples - the whole plate is analyzed with a single run. Standard fragment analysis run uses G5 dye set (6-FAM, VIC, NED PET with LIZ size standard). With a standard setting for a Fragment Analysis Run a resolution of 1 bp is achieved up to 500 bp with GeneScan-500 LIZ size standard. Please contact SeqLab staff if you wish to use other dyes, other size standards or fragments longer than 500 bp.
- Click here to learn about fluorescent dyes in fragment analysis.
- If you are setting up a new fragment analysis assay please read first Assay desingn / testing.
- Full 96 well plate ready for electrophoresis (PCR + Hi-Di + size standard). Each well must contain at least Hi-Di formamide. Use only ABI 96-well Optical reaction plates with barcodes (PN 4306737).
- Fragment analysis plate delivered to SeqLab fridge or by mail / courier >> Deliver samples.
- Completely filled fragment analysis submission form as e-mail attachment sent to fimm-seqlab(ät)helsinki.fi
Fragment Analysis run includes
- Electrophoresis with ABI3730xl DNA Analyzer
- Initial electrophoresis run quality assessment
- Storage of electrophoresis run as a ZIPped file into a database.
- Automatic e-mail notification when results are saved to database.
Fragment Analysis Run protocol
Prepare a master mix of Hi-Di Formamide and GeneScan LIZ-500 size standard (or other):
- Pipette 1 ml of Hi-Di formamide into a eppendorf tube.
- Add 2.5 µl of GeneScan LIZ-500 size standard into eppendorf tube.
- Vortex thoroughly.
- Pipette 10 µl into each well (also to wells without samples) of a MicroAmp 96-Well Reaction Plate (with barcode).
- Add 2 µl of diluted PCR products into each well.
- Spin samples down with plate centrifuge and seal properly.
NOTE: Keep the amount of size standard constant from assay to assay - it is enough for successful Fragment Analysis Run. Adjust the dilution of your PCR products in the sample if necessary to gain balanced PCR vs size standard ratio.
NOTE: Fill each well at least with Hi-Di Formamide. Empty well may lead to capillary damage when injecting with high voltage. If you have empty wells in the run, fill them with duplicates or other dilutions of your PCR fragments.