Strain-Dependent Variation in 18S Ribosomal DNA Copy Numbers in Aspergillus fumigatus
Strain-Dependent Variation in 18S Ribosomal DNA Copy Numbers in Aspergillus fumigatus
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DOI：
10.1128/jcm.02073-08
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发表时间：
2009-05-01
影响因子：
9.4
通讯作者：
Wickes, B. L.
中科院分区：
文献类型：
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作者：
Herrera, M. L.;Vallor, A. C.;Wickes, B. L.
计数烟曲霉CFU可能具有挑战性，因为通过平板计数确定CFU可能很困难。然而，通过定量实时PCR（qPCR）的CFU测定变得越来越普遍，并且通常依赖于检测多拷贝rRNA基因的亚基之一。本研究旨在确定不同的黑曲霉核糖体DNA拷贝数是恒定的还是可变的。烟曲霉菌株FKS 1用作单拷贝对照基因，并针对单拷贝（pyrG和ARG4）和多拷贝（arsC）对照进行验证。18S rDNA亚基的拷贝数，然后确定了各种分离株，并发现不同的菌株，从38至91个拷贝每个基因组。暴露于许多不同环境和生长条件后18S rDNA拷贝数的稳定性研究表明，拷贝数是稳定的，在所有条件下变化小于一个拷贝，包括从动物模型中回收的分离株。这些结果表明，虽然核糖体基因是qPCR计数的极好靶点，但在将其用作定量分析靶点之前应确定拷贝数。
Enumerating Aspergillus fumigatus CFU can be challenging since CFU determination by plate count can be difficult. CFU determination by quantitative real-time PCR (qPCR), however, is becoming increasingly common and usually relies on detecting one of the subunits of the multicopy rRNA genes. This study was undertaken to determine if ribosomal DNA (rDNA) copy number was constant or variable among different A. fumigatus isolates. FKS1 was used as a single-copy control gene and was validated against single-copy (pyrG and ARG4) and multicopy (arsC) controls. The copy numbers of the 18S rDNA subunit were then determined for a variety of isolates and were found to vary with the strain, from 38 to 91 copies per genome. Investigation of the stability of the 18S rDNA copy number after exposure to a number of different environmental and growth conditions revealed that the copy number was stable, varying less than one copy across all conditions, including in isolates recovered from an animal model. These results suggest that while the ribosomal genes are excellent targets for enumeration by qPCR, the copy number should be determined prior to using them as targets for quantitative analysis.