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The primer fI TS 9 prevents chimera formation during fungal DNA amplification in a bark beetle DNA background
Author(s) -
Strid Y.,
Ihrmark K.,
Stenlid J.
Publication year - 2015
Publication title -
forest pathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.535
H-Index - 49
eISSN - 1439-0329
pISSN - 1437-4781
DOI - 10.1111/efp.12156
Subject(s) - biology , primer (cosmetics) , dna , chimera (genetics) , dna sequencing , polymerase chain reaction , microbiology and biotechnology , ribosomal dna , genetics , gene , phylogenetics , chemistry , organic chemistry
Summary Insects are frequently associated with fungi in natural habitats. Molecular detection of fungal species using the primer pair ITS 1 F and ITS 4 to amplify the ribosomal ITS region has become standard for studies of fungal ecology. When addressing insect and fungal DNA together, sequencing of the ITS region often results in chimeras due to ITS 4 binding to both insect and fungal DNA . Using the newly developed primer f ITS 9, placed in the 5.8S region in the middle of the ITS region, we show that chimeras (insect–fungal, fungal–fungal, fungal–plants or fungal–mammals) can be avoided and a shorter and less variable DNA sequence product can be obtained. This method allows a more targeted amplification of fungal DNA from mixed samples and decreases the necessity for a nested PCR approach.