<pyrosequencingAssay>

This element defines a pyrosequencing assay. Pyrosequencing is a PCR-based genotyping assay allowing rapid high-throughput analysis of SNPs, insertions and deletions. The technology is based on sequencing by synthesis whereby the incorporation of a nucleotide into DNA is utilized in an enzyme cascade that ultimately emits light. Unincorporated nucleotides are removed from the system by apyrase eliminating background.

The result of a pyrosequencing assay is reported within a genotypingResult element.

The relationship between the primers and the reference sequence depends on the coordinate system that is used. The forward primer's annealing positions will always be less than the annealing positions of the reverse primer since the forward primer should fall to the logical left of the reverse primer in the coordinate system. This means that if the coordinate system is in the forward orientation and on the plus strand, then:

• the forward primer must anneal to the left of the reverse primer in the reference sequence, and the annealing portion of the sequence must match some substring of the reference sequence, and
• the reverse primer must anneal to the right of the forward primer in the reference sequence, and the annealing portion of the sequence must match some substring of the reverse compliment of the reference sequence.

If the coordinate system is in the reverse orientation and on the minus strand, just swap the forward and reverse primers in the statements above.

Note that it is an error for the coordinate system to be in the forward orientation and on the minus strand or in the reverse orientation and on the plus strand for pyrosequencing assays.

Attributes

pharmgkbId	A unique identifier assigned by the PharmGKB (the PharmGKB Accession Id). This attribute is used to refer to data that is already in the PharmGKB and is only supplied when editing or adding to such data. See the FAQ for details.
localId	A unique identifier for this element assigned by the submitter. The only rule it must follow is that no two XML elements can be assigned the same localId within a single XML document. See the FAQ for details.

note	A note.
submitter	This element's submitter.
interrogatedPosition	The position on the reference sequence analyzed by this assay.
analyzedRegion	The region of the reference sequence analyzed by this assay.
forwardPrimer	The forward sequencing primer that anneals 5' with respect to the reverse primer.
reversePrimer	The reverse sequencing primer that anneals 3' with respect to the forward primer.
pyrosequencingPrimer	The pyrosequencing primer that is used on the biotinylated strand.
method	The experimental method used to perform this assay.

Validation Rules

Content validation performed on this element:

• The interrogatedPosition must lie on the reference sequence.
• The start and stop positions of the analyzedRegion must lie on the reference sequence.

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