Abstract:
:High-quality RNA preparations are critical for further applications such as reverse transcriptase-polymerase chain reaction (RT-PCR) transcript amplifications, and elaboration of cDNA and expressed sequence tag libraries. Melanins are phenolic compounds present in many fungi and apparently play key roles in fungi pathogenesis and survival. However, during RNA extraction these compounds constitute a significant challenge to extraction of substantial quantities of high-quality RNA, and consequently to preparation of cDNA libraries. No method currently exists for RNA extraction from Mycosphaerella fijiensis that produces high quantities of melanin-free RNA. This fungus is the most important pathogen of cultivated Musa sp. varieties. A comparison is made between results obtained from the Trizol and RNeasy protocols for RNA extraction, two commercially available methods commonly used to obtain RNA from various sources. An improved methodology is described that allows isolation of intact RNA and elimination of melanins from M. fijiensis mycelium. RNA quality is evaluated by electrophoresis in formaldehyde-agarose gels, RT into cDNAs, and subsequent PCR amplification using primers designed against actin and beta- tubulin from fungi.
journal_name
Mol Biotechnoljournal_title
Molecular biotechnologyauthors
Islas-Flores I,Peraza-Echeverría L,Canto-Canché B,Rodríguez-García CMdoi
10.1385/MB:34:1:45subject
Has Abstractpub_date
2006-09-01 00:00:00pages
45-50issue
1eissn
1073-6085issn
1559-0305pii
MB:34:1:45journal_volume
34pub_type
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