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Abstract Detail

Mycological Section

Hughes, Karen Woodbury [1].

The consequences of cloning: clone chimeras, their frequency and distribution.

Cloning of PCR products has been used in environmental sampling and to resolve contributing haplotypes in heterozygotes. Unfortunately, the PCR/cloning process may generate chimeric sequences, which are thought to occur which an incomplete transcript is used as a primer in the next round or amplification and/or if polymerase template switching occurs. Hybrids between two nrITS2 genotypes of Gymnopus dichrous are common in nature and cloning was used to resolve individual haplotypes but clone chimeras were frequently recovered. Because nrITS2 homozygotes for haplotypes DI and DII exist in nature, chimeras were identifiable. The frequency of chimeric sequences was 16%. Template switching was non-random. Examination of potential DNA folding during the annealing portion of the PCR process indicated that regions of potential secondary structure were not associated with template switching leading to chimeric sequences. Chimeric sequences exist in GenBank and may create problems with fungal barcode identifications.

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1 - University of Tennessee, Ecology and Evolutionary Biology, 300 Hesler Biology Bldg, Knoxville, TN, 37996, USA

clone chimera

Presentation Type: Poster:Posters for Sections
Session: P
Location: Hall D/The Shaw Conference Centre
Date: Monday, July 27th, 2015
Time: 5:30 PM
Number: PMY065
Abstract ID:413
Candidate for Awards:None

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