Questions tagged [primer]

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Hard-clip primers in a bam file

I am working on a sample which was sequenced using an amplicon kit and I would like to ensure that no variants are called in the PCR primer sequences. I know that the primer sequences are part of the ...
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1answer
43 views

Directed mutagenesis primer design by back to back PCR of constructs

We're mutating a virus construct (virus-like particle [VLP]) using back to back primer PCR for a VLP construct and want to optimise the primer design. A regular primer design program (Primer3) isn't ...
1
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1answer
115 views

Hits in Primer-BLAST not found with programmatic blastn query

Summary: I'm writing an automated tool for designing Sanger sequencing primers (other tools don't do exactly what I want) that will be incorporated in a production diagnostic laboratory for on-demand ...
3
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1answer
79 views

programmatic secondary structure prediction for >36-mer DNA oligonucleotides

I'm writing a tool to automate Sanger sequencing primer design for a production lab that uses a universal-tail chemistry Sanger sequencing to verify NGS results. Essentially, the template DNA is ...
2
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1answer
76 views

What is the “primer masterfile” format?

I am working with an amplicon kit from Swift Bioscience and they provide the primer locations in what they call a "masterfile". The format is like this: ...
2
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1answer
95 views

Aligning sequence and comparing it against primer

I am looking to show how a primer is consistent among some genomic data. I have a primer of about 23bp and looking to compare it to about 5000 genomic sequences of 10kb. I am unsure how to format it ...
1
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0answers
30 views

Which software is recommended for sequencing primers selection?

I have a list of primers for sequencing fungal ITS1/2 regions. I also have access to different fungal databases. I would like to perform in silico PCR analysis for comparing different primer pairs and ...
2
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1answer
505 views

What is a poly(T) primer anchor sequence?

I am reading Tang et al. (2009). In the method description it is stated that: A single cell is manually picked under a microscope and lysed. Then mRNAs are reverse-transcribed into cDNAs using a ...