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The sequencing facility

Service overview

DNA sequencing

 

Automated Sanger DNA Sequencing is performed using capillary-based Applied Biosystem 3730XL Analyzer. The instrument offers high throughput and sensitivity. Samples of single stranded, double stranded DNA, and PCR products, can be sequenced with reads >800 base pairs.

 

The sequencing PCR is performed by the user with high-quality template DNA, 1 primer, BigDye terminator 3.1 mix and sequencing buffer.

The clean-up of DNA at the sequencing lab is robotized.

Sequences are usually available overnight on our server.                                                        

 

Fragment analysis

Applications:

    Micro satellite Analysis

    AFLP Analysis (amplified fragment length polymorphisms)

    Gene Expression Profiling

    Mutation Detection

 

Fluorescently labeled fragments are extracted at the Sequencing Lab and detected using the Applied Biosystems 3730XL Analyzer and then interpreted using the GeneMapper or GeneMarker analysis software.  Five different colored fluorescent dyes can be detected in one sample.  One of the dye colors is used for a labeled size standard that is added to each lane to allow comparison of samples from lane to lane or gel to gel.  The analysis software uses the size standard to create a standard curve for each lane and then determines the length of each dye-labeled fragment by comparing it with the standard curve for that specific lane.  Accuracy of less than one base difference is seen between sample replicates.  Higher throughput can be achieved by multiplexing different dyes and fragment sizes together.