The DOI matches its Crossref record; 2 of the 2 references it cites were found in a registry or an index.
Primer3—new capabilities and interfaces
Open accessAbstract
Polymerase chain reaction (PCR) is a basic molecular biology technique with a multiplicity of uses, including deoxyribonucleic acid cloning and sequencing, functional analysis of genes, diagnosis of diseases, genotyping and discovery of genetic variants. Reliable primer design is crucial for successful PCR, and for over a decade, the open-source Primer3 software has been widely used for primer design, often in high-throughput genomics applications. It has also been incorporated into numerous publicly available software packages and web services. During this period, we have greatly expanded Primer3's functionality. In this article, we describe Primer3's current capabilities, emphasizing recent improvements. The most notable enhancements incorporate more accurate thermodynamic models in the primer design process, both to improve melting temperature prediction and to reduce the likelihood that primers will form hairpins or dimers. Additional enhancements include more precise control of primer placement-a change motivated partly by opportunities to use whole-genome sequences to improve primer specificity. We also added features to increase ease of use, including the ability to save and re-use parameter settings and the ability to require that individual primers not be used in more than one primer pair. We have made the core code more modular and provided cleaner programming interfaces to further ease integration with other software. These improvements position Primer3 for continued use with genome-scale data in the decade ahead.
Overview
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Integrity signals
Registry
Cites
Statistics
Verification
Claims the paper makes
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
The self-check needs the paper’s full text.
Citations
Cites
Ultrafast and memory-efficient alignment of short DNA sequences to the human genome.
Langmead, B., Trapnell, C., Pop, M., & Salzberg, S. L. (2009) · Genome Biology · doi:10.1186/gb-2009-10-3-r25 · Cited by 21,849
Thermodynamics and NMR of Internal G·T Mismatches in DNA
Allawi, H. T., & SantaLucia, J. (1997) · Biochemistry · doi:10.1021/bi962590c · Cited by 623 · cited 2×
“The relevant parameters are taken from the following: (13–20).”
Background
Cited by
Personalized RNA mutanome vaccines mobilize poly-specific therapeutic immunity against cancer
Sahin, U., Derhovanessian, E., Miller, M., Kloke, B.-P., Simon, P., Löwer, M., Bukur, V., Tadmor, A. D., Luxemburger, U., Schrörs, B., et al. (2017) · Nature · doi:10.1038/nature23003 · Cited by 2,488
YTHDF3 facilitates translation and decay of N6-methyladenosine-modified RNA
Shi, H., Wang, X., Lu, Z., Zhao, B. S., Ma, H., Hsu, P. J., Liu, C., & He, C. (2017) · Cell Research · doi:10.1038/cr.2017.15 · Cited by 1,815
Transcriptome-wide isoform-level dysregulation in ASD, schizophrenia, and bipolar disorder
Gandal, M. J., Zhang, P., Hadjimichael, E., Walker, R. L., Chen, C., Liu, S., Won, H., van Bakel, H., Varghese, M., Wang, Y., et al. (2018) · Science · doi:10.1126/science.aat8127 · Cited by 1,247
Actively personalized vaccination trial for newly diagnosed glioblastoma
Hilf, N., Kuttruff-Coqui, S., Frenzel, K., Bukur, V., Stevanović, S., Gouttefangeas, C., Platten, M., Tabatabai, G., Dutoit, V., van der Burg, S. H., et al. (2019) · Nature · doi:10.1038/s41586-018-0810-y · Cited by 938
Predicting and Manipulating Cardiac Drug Inactivation by the Human Gut Bacterium Eggerthella lenta
Haiser, H. J., Gootenberg, D. B., Chatman, K., Sirasani, G., Balskus, E. P., & Turnbaugh, P. J. (2013) · Science · doi:10.1126/science.1235872 · Cited by 704
Genomic profiling of DNA methyltransferases reveals a role for DNMT3B in genic methylation
Baubec, T., Colombo, D. F., Wirbelauer, C., Schmidt, J., Burger, L., Krebs, A. R., Akalin, A., & Schübeler, D. (2015) · Nature · doi:10.1038/nature14176 · Cited by 705
Accurate and efficient detection of gene fusions from RNA sequencing data
Uhrig, S., Ellermann, J., Walther, T., Burkhardt, P., Fröhlich, M., Hutter, B., Toprak, U. H., Neumann, O., Stenzinger, A., Scholl, C., et al. (2021) · Genome Research · doi:10.1101/gr.257246.119 · Cited by 526
MAFG-driven astrocytes promote CNS inflammation
Wheeler, M. A., Clark, I. C., Tjon, E. C., Li, Z., Zandee, S. E. J., Couturier, C. P., Watson, B. R., Scalisi, G., Alkwai, S., Rothhammer, V., et al. (2020) · Nature · doi:10.1038/s41586-020-1999-0 · Cited by 501
DNA Repair Profiling Reveals Nonrandom Outcomes at Cas9-Mediated Breaks
van Overbeek, M., Capurso, D., Carter, M. M., Thompson, M. S., Frias, E., Russ, C., Reece-Hoyes, J. S., Nye, C., Gradia, S., Vidal, B., et al. (2016) · Molecular Cell · doi:10.1016/j.molcel.2016.06.037 · Cited by 454
Hepatocytes direct the formation of a pro-metastatic niche in the liver
Lee, J. W., Stone, M. L., Porrett, P. M., Thomas, S. K., Komar, C. A., Li, J. H., Delman, D., Graham, K., Gladney, W. L., Hua, X., et al. (2019) · Nature · doi:10.1038/s41586-019-1004-y · Cited by 402
A SWI/SNF-related autism syndrome caused by de novo mutations in ADNP
Helsmoortel, C., Vulto-van Silfhout, A. T., Coe, B. P., Vandeweyer, G., Rooms, L., van den Ende, J., Schuurs-Hoeijmakers, J. H. M., Marcelis, C. L., Willemsen, M. H., Vissers, L. E. L. M., et al. (2014) · Nature Genetics · doi:10.1038/ng.2899 · Cited by 354
Highly rearranged chromosomes reveal uncoupling between genome topology and gene expression
Ghavi-Helm, Y., Jankowski, A., Meiers, S., Viales, R. R., Korbel, J. O., & Furlong, E. E. M. (2019) · Nature Genetics · doi:10.1038/s41588-019-0462-3 · Cited by 354
Gut-licensed IFNγ+ NK cells drive LAMP1+TRAIL+ anti-inflammatory astrocytes
Sanmarco, L. M., Wheeler, M. A., Gutiérrez-Vázquez, C., Polonio, C. M., Linnerbauer, M., Pinho-Ribeiro, F. A., Li, Z., Giovannoni, F., Batterman, K. V., Scalisi, G., et al. (2021) · Nature · doi:10.1038/s41586-020-03116-4 · Cited by 331
Montague, M. J., Li, G., Gandolfi, B., Khan, R., Aken, B. L., Searle, S. M. J., Minx, P., Hillier, L. W., Koboldt, D. C., Davis, B. W., et al. (2014) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1410083111 · Cited by 340
Untangling the brain’s neuroinflammatory and neurodegenerative transcriptional responses
Srinivasan, K., Friedman, B. A., Larson, J. L., Lauffer, B. E., Goldstein, L. D., Appling, L. L., Borneo, J., Poon, C., Ho, T., Cai, F., et al. (2016) · Nature Communications · doi:10.1038/ncomms11295 · Cited by 336
Highly multiplexed simultaneous detection of RNAs and proteins in single cells
Frei, A. P., Bava, F.-A., Zunder, E. R., Hsieh, E. W. Y., Chen, S.-Y., Nolan, G. P., & Gherardini, P. F. (2016) · Nature Methods · doi:10.1038/nmeth.3742 · Cited by 330
Moore, M. J., Scheel, T. K. H., Luna, J. M., Park, C. Y., Fak, J. J., Nishiuchi, E., Rice, C. M., & Darnell, R. B. (2015) · Nature Communications · doi:10.1038/ncomms9864 · Cited by 326
Genome-wide de novo risk score implicates promoter variation in autism spectrum disorder
An, J.-Y., Lin, K., Zhu, L., Werling, D. M., Dong, S., Brand, H., Wang, H. Z., Zhao, X., Schwartz, G. B., Collins, R. L., et al. (2018) · Science · doi:10.1126/science.aat6576 · Cited by 316
Walking along chromosomes with super-resolution imaging, contact maps, and integrative modeling
Nir, G., Farabella, I., Pérez Estrada, C., Ebeling, C. G., Beliveau, B. J., Sasaki, H. M., Lee, S. D., Nguyen, S. C., McCole, R. B., Chattoraj, S., et al. (2018) · PLOS Genetics · doi:10.1371/journal.pgen.1007872 · Cited by 261
GC content shapes mRNA storage and decay in human cells
Courel, M., Clément, Y., Bossevain, C., Foretek, D., Vidal Cruchez, O., Yi, Z., Bénard, M., Benassy, M.-N., Kress, M., Vindry, C., et al. (2019) · eLife · doi:10.7554/elife.49708 · Cited by 250
Targeted Perturb-seq enables genome-scale genetic screens in single cells
Schraivogel, D., Gschwind, A. R., Milbank, J. H., Leonce, D. R., Jakob, P., Mathur, L., Korbel, J. O., Merten, C. A., Velten, L., & Steinmetz, L. M. (2020) · Nature Methods · doi:10.1038/s41592-020-0837-5 · Cited by 245
Persistence of Supplemented Bifidobacterium longum subsp. infantis EVC001 in Breastfed Infants
Frese, S. A., Hutton, A. A., Contreras, L. N., Shaw, C. A., Palumbo, M. C., Casaburi, G., Xu, G., Davis, J. C. C., Lebrilla, C. B., Henrick, B. M., et al. (2017) · mSphere · doi:10.1128/msphere.00501-17 · Cited by 231
Couraud, S., Vaca-Paniagua, F., Villar, S., Oliver, J., Schuster, T., Blanché, H., Girard, N., Trédaniel, J., Guilleminault, L., Gervais, R., et al. (2014) · Clinical Cancer Research · doi:10.1158/1078-0432.ccr-13-3063 · Cited by 215
Dietary fat and gut microbiota interactions determine diet-induced obesity in mice
Kübeck, R., Bonet-Ripoll, C., Hoffmann, C., Walker, A., Müller, V. M., Schüppel, V. L., Lagkouvardos, I., Scholz, B., Engel, K.-H., Daniel, H., et al. (2016) · Molecular Metabolism · doi:10.1016/j.molmet.2016.10.001 · Cited by 211
L1-associated genomic regions are deleted in somatic cells of the healthy human brain
Erwin, J. A., Paquola, A. C. M., Singer, T., Gallina, I., Novotny, M., Quayle, C., Bedrosian, T. A., Alves, F. I. A., Butcher, C. R., Herdy, J. R., et al. (2016) · Nature Neuroscience · doi:10.1038/nn.4388 · Cited by 195
Kazachenka, A., Bertozzi, T. M., Sjoberg-Herrera, M. K., Walker, N., Gardner, J., Gunning, R., Pahita, E., Adams, S., Adams, D., & Ferguson-Smith, A. C. (2018) · Cell · doi:10.1016/j.cell.2018.09.043 · Cited by 181
Roager, H. M., Licht, T. R., Poulsen, S. K., Larsen, T. M., & Bahl, M. I. (2014) · Applied and Environmental Microbiology · doi:10.1128/aem.03549-13 · Cited by 194
vonHoldt, B. M., Shuldiner, E., Koch, I. J., Kartzinel, R. Y., Hogan, A., Brubaker, L., Wanser, S., Stahler, D., Wynne, C. D. L., Ostrander, E. A., et al. (2017) · Science Advances · doi:10.1126/sciadv.1700398 · Cited by 186
Pendleton, A. L., Shen, F., Taravella, A. M., Emery, S., Veeramah, K. R., Boyko, A. R., & Kidd, J. M. (2018) · BMC Biology · doi:10.1186/s12915-018-0535-2 · Cited by 156
ATG16L1 orchestrates interleukin-22 signaling in the intestinal epithelium via cGAS–STING
Aden, K., Tran, F., Ito, G., Sheibani-Tezerji, R., Lipinski, S., Kuiper, J. W., Tschurtschenthaler, M., Saveljeva, S., Bhattacharyya, J., Häsler, R., et al. (2018) · Journal of Experimental Medicine · doi:10.1084/jem.20171029 · Cited by 167
Howrigan, D. P., Rose, S. A., Samocha, K. E., Fromer, M., Cerrato, F., Chen, W. J., Churchhouse, C., Chambert, K., Chandler, S. D., Daly, M. J., et al. (2020) · Nature Neuroscience · doi:10.1038/s41593-019-0564-3 · Cited by 168
Adams, J. M., Pei, H., Sandoval, D. A., Seeley, R. J., Chang, R. B., Liberles, S. D., & Olson, D. P. (2018) · Diabetes · doi:10.2337/db17-1385 · Cited by 159
Clostridium butyricum Strains and Dysbiosis Linked to Necrotizing Enterocolitis in Preterm Neonates
Cassir, N., Benamar, S., Khalil, J. B., Croce, O., Saint-Faust, M., Jacquot, A., Million, M., Azza, S., Armstrong, N., Henry, M., et al. (2015) · Clinical Infectious Diseases · doi:10.1093/cid/civ468 · Cited by 151
Ilumäe, A.-M., Reidla, M., Chukhryaeva, M., Järve, M., Post, H., Karmin, M., Saag, L., Agdzhoyan, A., Kushniarevich, A., Litvinov, S., et al. (2016) · The American Journal of Human Genetics · doi:10.1016/j.ajhg.2016.05.025 · Cited by 134
A scalable platform for the development of cell-type-specific viral drivers
Hrvatin, S., Tzeng, C. P., Nagy, M. A., Stroud, H., Koutsioumpa, C., Wilcox, O. F., Assad, E. G., Green, J., Harvey, C. D., Griffith, E. C., et al. (2019) · eLife · doi:10.7554/elife.48089 · Cited by 118
Jakobsen, N. A., Turkalj, S., Zeng, A. G. X., Stoilova, B., Metzner, M., Rahmig, S., Nagree, M. S., Shah, S., Moore, R., Usukhbayar, B., et al. (2024) · Cell Stem Cell · doi:10.1016/j.stem.2024.05.010 · Cited by 130
The Solanum tuberosum GBSSI gene: a target for assessing gene and base editing in tetraploid potato
Veillet, F., Chauvin, L., Kermarrec, M.-P., Sevestre, F., Merrer, M., Terret, Z., Szydlowski, N., Devaux, P., Gallois, J.-L., & Chauvin, J.-E. (2019) · Plant Cell Reports · doi:10.1007/s00299-019-02426-w · Cited by 124
Thézé, J., Li, T., du Plessis, L., Bouquet, J., Kraemer, M. U. G., Somasekar, S., Yu, G., de Cesare, M., Balmaseda, A., Kuan, G., et al. (2018) · Cell Host & Microbe · doi:10.1016/j.chom.2018.04.017 · Cited by 106
Benefits of higher resistance-training volume are related to ribosome biogenesis
Hammarström, D., Øfsteng, S., Koll, L., Hanestadhaugen, M., Hollan, I., Apró, W., Whist, J. E., Blomstrand, E., Rønnestad, B. R., & Ellefsen, S. (2020) · The Journal of Physiology · doi:10.1113/jp278455 · Cited by 99
Spettel, K., Barousch, W., Makristathis, A., Zeller, I., Nehr, M., Selitsch, B., Lackner, M., Rath, P.-M., Steinmann, J., & Willinger, B. (2019) · PLoS ONE · doi:10.1371/journal.pone.0210397 · Cited by 86
Vieira, W. A. S., Lima, W. G., Nascimento, E. S., Michereff, S. J., Câmara, M. P. S., & Doyle, V. P. (2017) · Mycologia · doi:10.1080/00275514.2017.1418577 · Cited by 83
Denisovan, modern human and mouse TNFAIP3 alleles tune A20 phosphorylation and immunity
Zammit, N. W., Siggs, O. M., Gray, P. E., Horikawa, K., Langley, D. B., Walters, S. N., Daley, S. R., Loetsch, C., Warren, J., Yap, J. Y., et al. (2019) · Nature Immunology · doi:10.1038/s41590-019-0492-0 · Cited by 80
Recombination of repeat elements generates somatic complexity in human genomes
Pascarella, G., Hon, C. C., Hashimoto, K., Busch, A., Luginbühl, J., Parr, C., Hin Yip, W., Abe, K., Kratz, A., Bonetti, A., et al. (2022) · Cell · doi:10.1016/j.cell.2022.06.032 · Cited by 77
Sun, X., Bizhanova, A., Matheson, T. D., Yu, J., Zhu, L. J., & Kaufman, P. D. (2017) · Molecular and Cellular Biology · doi:10.1128/mcb.00569-16 · Cited by 76
Environmental DNA as an efficient tool for detecting invasive crayfishes in freshwater ponds
Mauvisseau, Q., Coignet, A., Delaunay, C., Pinet, F., Bouchon, D., & Souty-Grosset, C. (2018) · Hydrobiologia · doi:10.1007/s10750-017-3288-y · Cited by 60
Rhode, C., Badenhorst, R., Hull, K. L., Greenwood, M. P., Bester‐van der Merwe, A. E., Andere, A. A., Picard, C. J., & Richards, C. (2020) · Animal Genetics · doi:10.1111/age.12961 · Cited by 58
Genetic diversity of Anopheles stephensi in Ethiopia provides insight into patterns of spread
Carter, T. E., Yared, S., Getachew, D., Spear, J., Choi, S. H., Samake, J. N., Mumba, P., Dengela, D., Yohannes, G., Chibsa, S., et al. (2021) · Parasites & Vectors · doi:10.1186/s13071-021-05097-3 · Cited by 48
Helliwell, K. E., Pandhal, J., Cooper, M. B., Longworth, J., Kudahl, U. J., Russo, D. A., Tomsett, E. V., Bunbury, F., Salmon, D. L., Smirnoff, N., et al. (2018) · New Phytologist · doi:10.1111/nph.14832 · Cited by 48
Clarke, C. M., Fok, V. T., Gustafson, J. A., Smyth, M. D., Timms, A. E., Frazar, C. D., Smith, J. D., Birgfeld, C. B., Lee, A., Ellenbogen, R. G., et al. (2018) · American Journal of Medical Genetics Part A · doi:10.1002/ajmg.a.38540 · Cited by 44
An ABCA4 loss-of-function mutation causes a canine form of Stargardt disease
Mäkeläinen, S., Gòdia, M., Hellsand, M., Viluma, A., Hahn, D., Makdoumi, K., Zeiss, C. J., Mellersh, C., Ricketts, S. L., Narfström, K., et al. (2019) · PLOS Genetics · doi:10.1371/journal.pgen.1007873 · Cited by 41
Banerjee, A. R., Kim, Y. J., & Kim, T. H. (2014) · Nucleic Acids Research · doi:10.1093/nar/gku1018 · Cited by 39
vonHoldt, B. M., Ji, S. S., Aardema, M. L., Stahler, D. R., Udell, M. A. R., & Sinsheimer, J. S. (2018) · Genome Biology and Evolution · doi:10.1093/gbe/evy112 · Cited by 31
Ray-Jones, H., Duffus, K., McGovern, A., Martin, P., Shi, C., Hankinson, J., Gough, O., Yarwood, A., Morris, A. P., Adamson, A., et al. (2020) · BMC Biology · doi:10.1186/s12915-020-00779-3 · Cited by 31
Sullivan, M. J., Goh, K. G. K., & Ulett, G. C. (2021) · mSphere · doi:10.1128/msphere.00105-21 · Cited by 23
Structural basis for an early stage of the photosystem II repair cycle in Chlamydomonas reinhardtii
Li, A., You, T., Pang, X., Wang, Y., Tian, L., Li, X., & Liu, Z. (2024) · Nature Communications · doi:10.1038/s41467-024-49532-2 · Cited by 26
Liu, X., Gündel, B., Li, X., Liu, J., Wright, A., Löhr, M., Arvidsson, G., & Heuchel, R. (2021) · Translational Oncology · doi:10.1016/j.tranon.2021.101107 · Cited by 24
Balinova, N., Post, H., Kushniarevich, A., Flores, R., Karmin, M., Sahakyan, H., Reidla, M., Metspalu, E., Litvinov, S., Dzhaubermezov, M., et al. (2019) · European Journal of Human Genetics · doi:10.1038/s41431-019-0399-0 · Cited by 20
Somatic Mutations and Genome Stability Maintenance in Clonal Coral Colonies
López, E. H., & Palumbi, S. R. (2020) · Molecular Biology and Evolution · doi:10.1093/molbev/msz270 · Cited by 18
Beckwith, K. S., Ødegård-Fougner, Ø., Morero, N. R., Barton, C., Schueder, F., Tang, W., Alexander, S., Peters, J. M., Jungmann, R., Birney, E., et al. (2025) · Nature Communications · doi:10.1038/s41467-025-61689-y · Cited by 13
Morson, N., Molenda, O., Picott, K. J., Richardson, R. E., & Edwards, E. A. (2022) · FEMS Microbes · doi:10.1093/femsmc/xtac021 · Cited by 10
Alexander, R. D., Castillejo-Pons, P., Melzer, N., Alsaif, O., Strotmann, V. I., Stahl, Y., Seale, M., & Morris, P. C. (2020) · doi:10.1101/2020.08.10.244889 · Cited by 7
Function and evolution of the aquaporin IsAQP1 in the Lyme disease vector Ixodes scapularis
Tsujimoto, H., Metz, H. C., Smith, A. A., Sakamoto, J. M., Pal, U., & Rasgon, J. L. (2023) · Insect Molecular Biology · doi:10.1111/imb.12833 · Cited by 6
Cathepsin X is a conserved cell death protein involved in algal response to environmental stress
Mizrachi, A., Sadeh, M., Ben-Dor, S., Dym, O., Ku, C., Feldmesser, E., Zarfin, A., Brunson, J. K., Allen, A. E., Jinkerson, R. E., et al. (2025) · Current Biology · doi:10.1016/j.cub.2025.03.045 · Cited by 5
Manor, J., Jangam, S. V., Chung, H.-L., Bhagwat, P., Andrews, J., Chester, H., Kondo, S., Srivastav, S., Botas, J., Moser, A. B., et al. (2024) · doi:10.1101/2024.09.23.614586 · Cited by 2
Egli-Spichtig, D., Zhang, M. Y. H., Li, A., Pastor Arroyo, E. M., Hernando, N., Wagner, C. A., Chang, W., & Perwad, F. (2021) · Scientific Reports · doi:10.1038/s41598-021-84735-3 · Cited by 1
Metadata
- Authors
- Untergasser, A., Cutcutache, I., Koressaar, T., Ye, J., Faircloth, B. C., Remm, M., & Rozen, S. G.
- Venue
- Nucleic Acids Research
- Publisher
- Oxford University Press (OUP)
- Published
- 2012-06-21
- Volume
- 40
- Issue
- 15
- Pages
- e115-e115
- ISSN
- 0305-1048 (ISSN-L)
- 1362-4962
- OpenAlex
- W2162098634
- Cited by
- 9,461
- References
- 2
- FWCI
- 109.43
- Type
- journal article
- Language
- en
- Open access
- gold
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
Are you an author, or do you see a problem with this page? Request a review
Data sources
Bibliographic data: Semantic Scholar, OpenAlex, Crossref.
Contains information from the Semantic Scholar Academic Graph Datasets, made available under the ODC Attribution License (ODC-BY 1.0).
Semantic Scholar’s data is described in: Kinney, R., et al. (2023). The Semantic Scholar Open Data Platform. arXiv:2301.10140.
