Ex-Ex Primer: An experimentally validated tool for designing oligonucleotides spanning spliced nucleic acid regions from multiple species

A comparative study of existing junction-primer-designing software revealed many limitations among them. Hence, we developed a new computational program, Ex-Ex Primer, which offers many improved, user-friendly features, and reliably creates junction primers and probes. This online suite can also be...

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Bibliographic Details
Published in:	Journal of biotechnology Vol. 343; pp. 1 - 6
Main Authors:	Govindkumar, Balagannavar, Kavyashree, Basavaraju, Patel, Krishna, Sasidharan, Kalesh, Siva Arumugam, T., Thomas, Lijo, Praveena, B.K.G., Raksha, H.N., Menon, R., Acharya, K.K.
Format:	Journal Article
Language:	English
Published:	Netherlands Elsevier B.V 10-01-2022
Subjects:	Alternative splicing DNA Primers - genetics Exon-exon junctions Junction primers Nucleic Acids Oligonucleotides Polymerase Chain Reaction Primer designing Software Alternative splicing Junction primers Primer designing Exon-exon junctions
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Summary:	A comparative study of existing junction-primer-designing software revealed many limitations among them. Hence, we developed a new computational program, Ex-Ex Primer, which offers many improved, user-friendly features, and reliably creates junction primers and probes. This online suite can also be used to design primers/probes from other sites of nucleic acid recombination, insertion, deletion, or splicing, and regular probes/primers. The threshold for Tm difference between the complete junctional primer vs its partial sequence, which maps to one of the junctional regions, was changed based on an important observation made during the initial experimental validations. The tool is now thoroughly checked with RT-PCR and RT-qPCR experiments with more than 250 primer pairs over a few years. The junction-primer-designing features of the software are also better than other equivalent tools. Visualizing the exons and introns across transcripts, and enabling primer designing based on information from Ensembl, are some of the unique features of this tool. The primers suggested by the tool can be used to detect the expression of known transcripts, to test the existence of predicted DNA or RNA joints via hybridization-based techniques, or for validation and in silico analysis of RNA-Seq. URL: http://resource2.ibab.ac.in/exprimer/. •Visualizations of transcript variants and junctions for primer/probe designing.•Tm difference set between partially annealed vs completely annealed junctional primer.•250 primer pairs were tested to validate the utility of the tool.•The tool also supports the designing of regular primers and probes.•NCBI and Ensembl transcript information displayed for selected ids of Human, Mouse, and Rat species.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0168-1656 1873-4863
DOI:	10.1016/j.jbiotec.2021.10.009