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The mCherry-tagged IRE1 and ire1 (ΔKR/Δ-box) copies were integrated from pRS305 (Sikorski and Hieter, 1989) based constructs.
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Once these provirus copies are integrated, there is no way to remove them from the host's genome.
The resulting cDNA copies are integrated more or less randomly into the host genome and usually are transcriptionally silent, due to the lack of upstream promoter elements [14], [29].
However, the prevalent assumption has been that multiple transgene copies are integrated in a tandem head-to-tail array with only rare exceptions of inverted repeats [28], [29], [30].
If multiple copies are integrated in head-to-tail tandem configuration into a single locus of genome, Southern blot analysis reveals the expected size after digestion with the appropriate restriction enzyme as shown in Fig. 1A.
This effect seems to be more pronounced the more copies are integrated (Aw and Polizzi 2013).
This indicates multiple copies are integrated per cell when using higher titers.
The idea is that short regions of viral mRNAs are copied by host reverse transcriptase and the copies are integrated in the genome where a few can be utilized to produce small RNAs interfering with the respective virus (imRNAs).
The occurrence of transformation events with multiple T-DNA copies being integrated in independent genomic loci of triticale opens up the opportunity to generate transgenic segregants with reduced copy number.
A later copy was integrated onto the GomSpace GomX-2 satellite.
The SpR copy was integrated into the genome of the ire1Δ strain from a pRS304 (Sikorski and Hieter, 1989) plasmid derivative of pDEP005 (Pincus et al., 2010).
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