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Primer mixtures in RAPD analysis
Department of Genetics, Lund University.
Högskolan Kristianstad, Institutionen för matematik och naturvetenskap. (Biomedicin)
Department of Clinical Chemistry, Malmö University Hospital,. (Biomedicin)ORCID-id: 0000-0002-9355-3901
2000 (Engelska)Ingår i: Hereditas, ISSN 0018-0661, E-ISSN 1601-5223, Vol. 132, nr 3, s. 203-208Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

RAPD (random amplified polymorphic DNA) is a multiplex marker system that conventionally uses single-primer PCR to amplify random DNA fragments. Because of its multiplex nature, it is frequently used in bulked segregant analysis (BSA). In view of the very large numbers of markers BSA often requires, we investigated the use of mixtures of primers as a method of increasing the number of markers available. Theoretically, if a single-primer reaction produces x bands on average, an unrestrained PCR process using a primers should produce xa2 bands. Initially, we investigated mixtures containing from one to five primers. The average number of products increased slightly from the single-primer to the multiple-primer case, whereas it was rather constant for the different multi-primer combinations. This deviation from the theoretical expectations, which we attribute to the effects of competition, shows mixtures of more than two primers to be inefficient. The properties of two-primer mixtures in which the proportions of the two primers were varied were also investigated. The intensities of most of the products were influenced by the proportions of the primers used to create the mixture. A good fit was obtained to a model in which the average competitive ability of a band is directly proportional to the probability of randomly obtaining the band-producing primer combination from the pool of primers. Using two-primer mixtures, a(a-1)/2 different two-primer mixtures can be produced. A comparison of different schemes for constructing the two-primer mixtures indicates that the degree of resampling is similar for all schemes. In conclusion, the use of two-primer mixtures is a simple but very powerful strategy in BSA as it can generate an extremely large number of markers.

Ort, förlag, år, upplaga, sidor
2000. Vol. 132, nr 3, s. 203-208
Nationell ämneskategori
Biologiska vetenskaper
Identifikatorer
URN: urn:nbn:se:hkr:diva-8187DOI: 10.1111/j.1601-5223.2000.00203.xPubMedID: 11075515OAI: oai:DiVA.org:hkr-8187DiVA, id: diva2:424490
Tillgänglig från: 2011-06-17 Skapad: 2011-06-17 Senast uppdaterad: 2019-07-03Bibliografiskt granskad

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Lind-Halldén, ChristinaHalldén, Christer

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