Ty Element Retrotransposition in Saccharomyces cerevisia

一年来，我们在以下方面取得了进展。通过在Ty-less菌株中重新引入活性元件，研究了Ty1拷贝数的动态变化，在该菌株中，驻留元件因长末端重复(LTR-LTR)重组而丢失。重新填充的菌株表现出与Ty1插入相关的染色体大小变化，但不会在基因上分离。在已知影响Ty1逆转录转座的遗传和环境条件下，元件的得率和丢失率已经使用基因标记的参考元件来确定。在不同条件下获得的Ty1获得和丢失比率表明，拷贝数随着时间的推移而变化，通过改变逆转录转座的速度，导致在酵母中观察到不同的拷贝数。在更广泛的范围内，我们的结果表明，LTR逆转座子的获得和丢失可以是高度动态的，并有可能在较短的进化时间段内重塑基因组。与Dwight V.Nissley博士(SAIC，Frederick MD)合作，我们已经证明了由Ty1和来自HIV-1的逆转录酶基因组成的杂交元件是检测、监测和分离萌芽酵母中耐药逆转录酶的有用工具。这种灵敏的表型分析检测临床样本中的耐药变异，当变异仅占病毒总数的0.3%至1.0%时。此外，我们的检测方法可以检测已知和新的逆转录酶变异，并应有助于研究微小耐药变异的进化和临床意义。

Over the past year, we have made progress in the following areas. Ty1 copy number dynamics has been studied by reintroducing active elements into a Ty-less strain where resident elements had been lost by long terminal repeat (LTR-LTR) recombination. Repopulated strains exhibit alterations in chromosome size that are associated with Ty1 insertions, but do not become genetically isolated. The rates of element gain and loss under genetic and environmental conditions known to affect Ty1 retrotransposition have been determined using genetically tagged reference elements. Ty1 gain and loss ratios obtained under different conditions suggest that copy number oscillates over time by altering the rate of retrotransposition, resulting in the diverse copy numbers observed in Saccharomyces. On a broader scale, our results suggest that the gain and loss of LTR-retrotransposons can be highly dynamic and has the potential to remodel genomes over short evolutionary time periods. In collaboration with Dr. Dwight V. Nissley (SAIC, Frederick MD), we have shown that hybrid elements composed of Ty1 and the reverse transcriptase gene from HIV-1 are useful tools for detecting, monitoring, and isolating drug-resistant reverse transcriptases in budding yeast. This sensitive phenotypic assay detects drug-resistant variants in clinical samples when the variants comprise as little as 0.3 to 1.0% of the virus population. Furthermore, our assay can detect of both known and novel reverse transcriptase variants and should be useful in studies of the evolution and clinical significance of minor drug-resistant variants.