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Primer-blast tries to find target-specific primers by placing candidate primers on unique template regions that are not similar to other targets.
Go to the Primer BLAST submission form. Enter the target sequence in FASTA format or an accession number of an NCBI nucleotide sequence in the PCR Template section of the form. If the NCBI mRNA reference sequence accession number is used, the tool will automatically design primers that are specific to that splice variant. If one or both primer ...
Nov 28, 2023 · The Basic Local Alignment Search Tool (BLAST) finds regions of local similarity between sequences. The program compares nucleotide or protein sequences to sequence databases and calculates the statistical significance of matches.
Primer-BLAST can only find non-specific primers for your template since there are other targets that are similar or identical to your template. You may choose to re-search for specific primers by allowing some of the highly similar off-targets if they are irrelevant to your PCR experiment.
One needs to design primers that are complementary to the template region of DNA. They are synthesized chemically by joining nucleotides together. One must selectively block and unblock repeatedly the reactive groups on a nucleotide when adding a nucleotide one at a time.
To design your primers, you can use the Oligo Calc: Oligonucleotide Properties Calculator or the Eurofins genomics calculator or the NIH Primer-BLAST. The following are the three (3) best online primer design tools that you can use to design your primers for your PCR experiment:
Oct 14, 2022 · Primer-BLAST is a combination of Primer3, a popular primer designing tool, and NCBI BLAST . Primer-BLAST has two basic modes. It can: Design primers to specifically amplify a template sequence or a set of related templates from a background nucleic acid population (BLAST database).
