Nucleic acid probes and methods for detecting candida DNA cells in blood
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文摘
Provided is an isolated double-stranded nucleic acid consisting essentially of the nucleotide sequence defined in the Sequence Listing by SEQ ID NO:5. This is the C. albicans ITS2 sequence and includes a nucleic acid comprising a nucleotide sequence that is specific for Candida albicans. Further examples of an isolated double stranded nucleic acid of the present invention consist essentially of the nucleotide sequences defined in the Sequence Listing by SEQ ID NOs:6-9. These are the ITS2 sequences for C. parapsilosis, C. tropicalis, C. glabrata and C. krusei. A method of diagnosing systemic candidiasis in a subject is also provided. The method comprises the steps of: (a) collecting blood from the subject into tubes containing detergent, polypropylene glycol, sodium polyanetholesulfonate, and sodium ethylene diamine trademark tetraacetic acid; (b) lysing Candida cells using ZYMOLYASE trademark-100T with agitation; (c) extracting and precipitating the DNA from the lysed cells; (d) amplifying the precipitated DNA using universal fungal primer pairs derived from the internal transcribed spacer regions of the Candida ribosomal DNA; and (e) detecting amplified DNA from Candida by hybridizing the amplified DNA with a probe that selectively hybridizes with Candida DNA, the presence of amplified DNA indicating systemic candidiasis.
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