Wheelan Sarah J, Scheifele Lisa Z, Martínez-Murillo Francisco, Irizarry Rafael A, Boeke Jef D
High Throughput Biology Center and Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Proc Natl Acad Sci U S A. 2006 Nov 21;103(47):17632-7. doi: 10.1073/pnas.0605450103. Epub 2006 Nov 13.
Mobile elements are important components of our genomes, with diverse and significant effects on phenotype. Not only can transposons inactivate genes by direct disruption and shuffle the genome through recombination, they can also alter gene expression subtly or powerfully. Currently active transposons are highly polymorphic in host populations, including, among hundreds of others, L1 and Alu elements in humans and Ty1 elements in yeast. For this reason, we wished to develop a simple genome-wide method for identifying all transposons in any given sample. We have designed a transposon insertion site profiling chip (TIP-chip), a microarray intended for use as a high-throughput technique for mapping transposon insertions. By selectively amplifying transposon flanking regions and hybridizing them to the array, we can locate all transposons present in a sample. We have tested the TIP-chip extensively to map Ty1 retrotransposon insertions in yeast and have achieved excellent results in two laboratory strains as well as in evolved Ty1 high-copy strains. We are able to identify all of the theoretically detectable transposons in the FY2 lab strain, with essentially no false positives. In addition, we mapped many new transposon copies in the high-copy Ty1 strain and determined its Ty1 insertion pattern.
移动元件是我们基因组的重要组成部分,对表型具有多样且显著的影响。转座子不仅可以通过直接破坏使基因失活,并通过重组打乱基因组,还能或微妙或有力地改变基因表达。目前活跃的转座子在宿主群体中具有高度多态性,在人类中包括数百种其他类型,如L1和Alu元件,在酵母中则有Ty1元件。因此,我们希望开发一种简单的全基因组方法,用于识别任何给定样本中的所有转座子。我们设计了一种转座子插入位点分析芯片(TIP-chip),这是一种微阵列,旨在用作绘制转座子插入位点的高通量技术。通过选择性地扩增转座子侧翼区域并将它们与阵列杂交,我们可以定位样本中存在的所有转座子。我们已经对TIP-chip进行了广泛测试,以绘制酵母中Ty1逆转座子的插入位点,并且在两个实验室菌株以及进化后的Ty1高拷贝菌株中都取得了优异的结果。我们能够识别FY2实验室菌株中所有理论上可检测到的转座子,基本没有假阳性。此外,我们在高拷贝Ty1菌株中绘制了许多新的转座子拷贝,并确定了其Ty1插入模式。