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1.
Figure 5

Figure 5. From: Local Definition of Ty1 Target Preference by Long Terminal Repeats and Clustered tRNA Genes.

Distribution of de novo insertions within 650 bases upstream of the TSSs of tGly and tThr gene families and individual preferred tDNAs. (A) Insertions upstream of the tGly family. (B) Insertions upstream of the tThr family. (C) Insertions upstream of individual tDNAs that received more than 15 insertion events. (Bottom, left) Composite of insertions upstream of all tDNA targets that received 15 or less insertion events. Insertions are plotted as described in the legend to . Gray bars highlight the 36 bases (18 bases on each side) around each integration peak (–85, –170, –265, and –335).

Nurjana Bachman, et al. Genome Res. 2004 July;14(7):1232-1247.
2.
Figure 3

Figure 3. From: Local Definition of Ty1 Target Preference by Long Terminal Repeats and Clustered tRNA Genes.

Number of insertions upstream of each member of the tGly and tThr gene families. (A) Number of insertions upstream of each of the 16 genomic tGly genes. All tGly genes assayed had GCC as their anticodon. Roman numerals represent chromosome on which the copy exists; arabic numerals after the comma represent the copy number in cases where more than one tGly exists on the same chromosome. For example, tGly IV, 1, represents the first (5′-most on the Watson strand) of the two copies of tGly on chromosome IV. The name for this gene in the Saccharomyces genome database (http://www.yeastgenome.org/) is tG(GCC)D1. (B) Number of insertions upstream of each of the 11 genomic tThr genes. All tThr assayed had AGU as their anticodon and were named by the same conventions described for tGly.

Nurjana Bachman, et al. Genome Res. 2004 July;14(7):1232-1247.
3.
Figure 2

Figure 2. From: Local Definition of Ty1 Target Preference by Long Terminal Repeats and Clustered tRNA Genes.

The insertion pattern upstream of the tGly and tThr gene families is periodic and depends on the expression of the marked Ty1 element and the Ty1 integrase protein. Four primer pairs were used to identify Ty1 insertions upstream of tGly and tThr genes in both orientations. (A) PCR products from each primer combination resulting from galactose induction with the pVIT41 Ty1 donor plasmid, without induction or without the plasmid. (B) The periodic insertion pattern depends on the presence of wild-type Ty1 integrase and does not require the homologous recombination pathway. Inductions were completed in a Δrad52 strain. The pNB19 plasmid containing a mutation in the catalytic site of integrase (–) or pVIT41 (WT) were used as the source of Ty1.

Nurjana Bachman, et al. Genome Res. 2004 July;14(7):1232-1247.
4.
Figure 4

Figure 4. From: Local Definition of Ty1 Target Preference by Long Terminal Repeats and Clustered tRNA Genes.

Distribution of de novo Ty1 insertions and endogenous Ty retrotransposon sequences upstream of tDNA targets in the yeast genome. (A) Composite diagram of all sequenced de novo insertion events within 650 bases upstream of the TSS of 35 different genomic tDNAs (n = 803). Each line represents insertion events into a single base position. The frequency of insertion events into a position is shown vs. the distance upstream of the tDNA target. Insertions begin 65 bases upstream of the tDNA, shown at the far right, and distance increases leftward across the x-axis. Ty1 insertions amplified using the orientation 1 primer are shown above the x-axis; those amplified with the orientation 2 primer are shown below the line. (B) Position of endogenous Ty1, Ty2, and Ty4 LTRs in the yeast genome upstream of the mature 5′ processed end of the tDNA target.

Nurjana Bachman, et al. Genome Res. 2004 July;14(7):1232-1247.
5.
Figure 1

Figure 1. From: Local Definition of Ty1 Target Preference by Long Terminal Repeats and Clustered tRNA Genes.

The PCR assay for targeted integration upstream of tDNAs in S. cerevisiae. The pVIT41 donor plasmid () contains a unique oligonucleotide-binding site referred to as SSB. LTRs are shown as triangles, with the wide part of the triangle representing the U3 region. After induction on galactose medium to express the Ty1, the Ty1 transcript is translated, reverse transcribed, and the resulting cDNA is integrated into the yeast genome in either orientation. Insertion events are detected by using a primer complementary to the SSB in the Ty1 element in combination with a tDNA-specific primer. PCR products labeled with an asterisk (*) are >6 kb long and are not generated by the PCR conditions used in these experiments.

Nurjana Bachman, et al. Genome Res. 2004 July;14(7):1232-1247.
6.
Figure 6

Figure 6. From: Local Definition of Ty1 Target Preference by Long Terminal Repeats and Clustered tRNA Genes.

Comparison of features around a hot and cold tGly target and effects on integration frequency. (A) Schematic of features surrounding the most preferred tGly target, tGlyII (tG(GCC)B) from JB1217 and the least preferred target, tGlyIV (tG(GCC)D1) from BY4741. (B) Various features were added into several positions around tGlyIV. The tIle with 60 bases of upstream sequence and 46 bases of downstream sequence was amplified from JB1217 tGlyII and inserted 60 bases upstream, 25 bases downstream, or 299 bases downstream of BY4741 tGlyIV. The LTR downstream of tGlyII (68% identity to a query Ty1 LTR [http://www.public.iastate.edu/∼voytas/], Ty1 H3 5′ LTR) was inserted in both orientations either 299 or 25 bases downstream of tGly. Back-to-back LTRs (the first LTR is 68% identical and the second 87% identical to the reference LTR) were added at position 299. The LTR insertions were combined with insertion of tIle. When the 180 bases containing the tIle were added 25 bases downstream of tGly, the LTR shifts downstream by 180 bases, to 479 bases downstream of the tGlyIV. The same PCR primer was used to amplify insertions upstream of tGly in every strain, and its distance from tGlyIV shifts depending on the length of sequences inserted at the position 25 bases downstream of tGlyIV. Histogram showing amounts of PCR product generated from PCR of insertions upstream of tGlyIV in each strain after normalization to actin is shown at right. Histogram represents the mean of either eight or nine independent trials per strain (quantitation described in Methods section). Error bars, SEM. An asterisk (*) to the right of the bar indicates a statistically significant difference (P ≤ 0.01 by the Mann-Whitney U-test) in PCR product compared with the amount generated in the BY4741 strain. (C) Gels showing the products generated using the PCR assay of insertions upstream of tGlyIV. Two different transformants from every strain are shown. PCR of the actin gene was performed to control for amount of input genomic DNA.

Nurjana Bachman, et al. Genome Res. 2004 July;14(7):1232-1247.

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