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Figure 1 | Mobile DNA

Figure 1

From: Host co-factors of the retrovirus-like transposon Ty1

Figure 1

Modified synthetic genetic array screen for rhfΔ mutants. (A) Schematic of the Ty1his3AI element that is used to assay the frequency of retrotransposition. Ty1 long terminal repeats are represented as tripartite black and white boxes that flank internal region of the element. The internal region has two partially overlapping ORFs, gag and pol (green rectangles). As illustrated, the gag ORF begins in the 5′ LTR. The his3AI retrotransposition indicator gene is inserted in non-coding DNA between the end of pol and the beginning of the 3′ LTR. The his3AI gene consists of a HIS3 gene (red rectangles) in the opposite orientation to gag and pol. HIS3 is interrupted by insertion of an artificial intron (AI; blue rectangle) in an unspliceable orientation; however, AI is spliced from the Ty1his3AI transcript. When splicing occurs and the transcript is reverse transcribed, a Ty1HIS3 cDNA is produced, which, when integrated into the genome, confers a His+ phenotype. (B) Schematic of the genetic manipulations used to generate haploid progeny containing the Ty1his3AI element, the rtt101Δ or med1Δ mutation, and an orfΔ mutation. The query strain containing the rtt101Δ or med1Δ and the Ty1his3AI-MET15 allele was mated to each orfΔ::kanMX strain in the yeast ORF deletion library. Following induction of transposition by growth of cells in YEPD broth at 20°, progeny were plated on YEPD agar containing G418 to assess growth (not shown) and SC -His agar to measure retrotransposition. (C) The results of the Ty1his3AI retrotransposition assay on one SC -His plate of rtt101Δ: LEU2orfΔ:kanMX progeny. Cells that sustained a retrotransposition event give rise to His+ papillae, which were counted at each address. Addresses that are blank lack progeny because of synthetic lethality or slow growth (green circles). Addresses with ≤5 His+ papillae (red circles) harbor progeny with reduced retrotransposition. The parental rtt101Δ strain (blue circle) was plated in an empty address prior to induction of retrotransposition.

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