Questions tagged [primer]

Short sequence of DNA which hybridises to a locus within a genome and is used to initate replication of a unit of DNA through the DNA polymerase enzyme. Essential for PCR

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Do you know a program/script to count how many sequences have mismatches on each position of my primers?

I'm working with primer analysis and design, and so far I've performing it in Excel but: a) I don't like Excel, b) I do it manually, c) I don't like Excel, d) it gets tedious when working with ...
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Can you give me a detailed explanation about PrimeBlast?

I am trying to write a research paper on PRIMER-BLAST. I have read something about it on https://www.ncbi.nlm.nih.gov/pubmed/22708584/ but still, it is not enough. So can you please tell me more ...
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Primer-Blast Custom Database [error]

I'm using Primer-Blast to compare a PCR template against a custom database of reference sequences, using Primer-Blast's Custom database functionality. However whenever I try to upload a FASTA file as ...
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96 views

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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54 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 ...
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1answer
155 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 ...
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1answer
86 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 ...
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107 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: ...
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1answer
123 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 ...
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31 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 ...
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778 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 ...