Pyrosequencing software

Open the Pyrosequencing software. A user name and password is required. This is usually set up with instrument installation. Individual or group-wide passwords can be used. 

Select the Batch run icon on the far left of the Pyrosequencing software, on the menu tree right click over batch runs and select new batch run. One to ten plates can be run in each batch. A unique name for each batch must be provided, and the instrument parameters must be selected for each batch. If barcoded plates are not used, uncheck the barcode field. 

Any primer design software, freely available or custom purchased may be used to design PCR primers for Pyrosequencing. The polymorphism may be in any position of the PCR amplicon from 1 base in from the 3 end of the PCR primer sequence to centered between the primers. SNPs, indels, repeats, etc., do not require specific PCR primer design modifications. 

Problem template loops will also be flagged in the Pyrosequencing primer design software. 

If an appropriate Pyrosequencing primer cannot be found as the critical scoring parameters are flagged, it is possible to trick the software to improve the search. As the software will only look five bases to either side of the SNP for a suitable... 

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