ABI Prism® 7900HT Sequence Detection System ... - OpenWetWare

Allelic
Discrimination
Checklist
Dissociation
(Melting) Curve
Checklist
4-4 Run Setup and Basic Operation
To create and set up a plate document for an allelic discrimination run:
Done Step Description See Page
1 Create an allelic discrimination plate document. 4-6
2a a. Create detectors for the allelic discrimination probes. 4-7
b. Create a marker for each allelic discrimination probe
pairing.
4-9
c. Copy the marker(s) to the plate document. 4-10
3a Assign detector tasks to the wells of the plate document
(NTC and Unknown).
4-11
4 If you would like to perform thermal cycling of the allelic
discrimination plate on the 7900HT instrument, create a
real-time plate document for the plate and program it with the
the method for the allelic discrimination run. Otherwise,
continue to step 5.
4-13
5 Choose from the following:
If running… Then…
a single plate continue to step 7.
the first plate in a series Save the plate document as an ABI PRISM
of plates with identical SDS Template Document as explained on
assay configurations page 4-17.
6 Create a plate document from the template created in step 5. 4-18
7 Configure the document with sample names and plate
information.
4-19
8 a. Prepare the allelic discrimination plate or plates and
perform thermal cycling on a designated thermal cycler.
b. Run the allelic discrimination plate or plates on the
7900HT instrument.
4-20
9 Analyze the run data. 5-3
a. Steps 2 and 3 can be eliminated by importing the plate document setup information from a tab-delimited
text file. See “Importing Plate Document Setup Table Files” on page B-2 for more information.
A dissociation curve analysis is performed as part of a real-time PCR run (absolute
quantification). To perform a dissociation curve, construct a plate document for
absolute quantification as explained on page 4-3 and configure the method with a
temperature ramp as explained on page 4-16.