What is the difference between BSP and MSP primers?

BSP primers cover no CpG, so one pair amplifies both alleles and the methylation state is read from the sequencing trace at every CpG in the product. MSP primers deliberately cover CpG sites near their 3-prime end, so each pair only amplifies one allele and the band itself is the result.

Why do the primer sequences look nothing like my sequence?

They are the converted oligos you would actually order. Bisulfite turns unmethylated cytosine into uracil, read as T, so a treated template is strongly T-rich and its two strands are no longer complementary.

Which CpG island definition does it use?

Either the classical Gardiner-Garden and Frommer criteria, as EMBOSS cpgplot encodes them, or the stricter Takai and Jones 2002 thresholds for promoter-associated islands. Both are selectable and every threshold is shown.

Amplify · experimental

Bisulfite methylation primers

Find the CpG islands in a sequence and design the primers that read methylation in one: a BSP pair that amplifies both alleles for sequencing, or the two MSP pairs whose bands are themselves the answer. Islands are called under published criteria you can see and change, primers are designed on the bisulfite-converted sequence, and the rule that actually protects the assay — enough non-CpG cytosines that a primer cannot bind untreated DNA — is enforced and counted per oligo.

Open Primer Designer Open the Primer test sample

Real sequences (GenBank plasmid, FASTA, and a protein family) for the Primer Designer

What it does

01CpG island prediction under two published criteria sets, every threshold adjustable
02In-silico bisulfite conversion of both alleles on both strands
03BSP primer pairs that cover no CpG, so both alleles amplify equally
04MSP primer pairs for the methylated and unmethylated allele in one run
05Non-CpG cytosine count per primer, the check that a primer cannot bind untreated DNA
06The island, the product and both primers drawn on the sequence map

What it runs on

CpG island prediction is validated base for base against EMBOSS cpgplot on a real sequence corpus; primer rules follow the criteria published for MethPrimer and MSP-HTPrimer, over the same golden-validated melting-temperature engine the other primer tools use. The panel runs on @ports/enzymes, @ports/oligo-structure, @ports/primer.

Everything runs client-side. Files are parsed in the browser and projects are saved on this machine, so an unpublished sequence is never uploaded.

Questions

What is the difference between BSP and MSP primers?: BSP primers cover no CpG, so one pair amplifies both alleles and the methylation state is read from the sequencing trace at every CpG in the product. MSP primers deliberately cover CpG sites near their 3-prime end, so each pair only amplifies one allele and the band itself is the result.
Why do the primer sequences look nothing like my sequence?: They are the converted oligos you would actually order. Bisulfite turns unmethylated cytosine into uracil, read as T, so a treated template is strongly T-rich and its two strands are no longer complementary.
Which CpG island definition does it use?: Either the classical Gardiner-Garden and Frommer criteria, as EMBOSS cpgplot encodes them, or the stricter Takai and Jones 2002 thresholds for promoter-associated islands. Both are selectable and every threshold is shown.

Also searched as

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