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In the third step, I-SceI-mediated DSB resulted in 27/36 (75%) of the colonies showing precise removal of the cassette, resulting in a clean insertion [Figure 3B (c)].
Staged alignment of high throughput sequencing data, where reads are first aligned to the transgene sequence and then only orphaned mates of those reads are aligned to the host genome, simplifies the analysis such that a single cluster of mapped reads indicates a clean insertion, whereas two clusters in close proximity can indicate a deletion in the host genome.
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The patient must receive adequate training on how to properly clean screw insertion sites and the external frame [6].
This success resulted from using an experimental design that primarily targeted the exponential phase and from the use of a clean genome without any insertion sequences [ 43] that could have masked transcriptional change patterns.
We developed a procedure (HangOut) that, for a single domain with specified insertion position, cleans erroneously extended PSI-BLAST alignments to generate better profiles.
The insertion area was cleaned with povidone, and the sterile cover (Cathereeplus™, Nichiban CO. Ltd., Tokyo, Japan) was applied to the probe.
Before post insertion, each post was cleaned with ethanol (70% vol).
Indeed, the presence of a NumtS inside a gene may cause loss of function, so selection may act to clean out the genome from disrupting NumtS insertion events.
It is important to note that most insertions are not the desired clean homologous recombination event.
If the first attempt failed, another attempt involved a second insertion with the original but cleaned NGT by the same procedure.
Following catheter insertion, the fistula site is cleaned and the catheter changed once weekly at GFC. Fistula closure is assessed using the dye test.
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