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Figure 2 | Genome Integrity

Figure 2

From: Pathway choice in DNA double strand break repair: observations of a balancing act

Figure 2

NHEJ repair assays. a) Linear plasmid DNA with 6 bp repeats at the ends is joined after transfection. The joints are amplified by PCR and digested using BstXI to distinguish between direct repair and microhomology mediated repair [21]. b) Repair of linearized plasmid DNA results in restoration of GFP expression. c) Cleavage by I-SceI and subsequent repair lead to loss of the middle splice donor and acceptor sites (SD and SA) and the adenoviral exon (AD), resulting in the expression of active GFP [22]. d) H2Kd fused to CD8 is expressed from the intact substrate. Repair of the oppositely oriented I-SceI breaks results in loss of H2Kd-CD8 and allows expression of CD4 [23]. e) Similar to d), the intact substrate expresses GFP, while the repaired substrate allows expression of RFP and loses GFP expression [24]. f) Between the opposite I-SceI sites, a translation start site is located, preventing translation of the XHATM resistance gene. Repair of the I-SceI breaks and loss of the intervening ATG results in XHATM resistance. The sequence around the breaks can be sequenced to monitor loss of nucleotides [25]. g) V(D)J recombination assay. Cleavage by the Rag1/2 endonuclease at the recombination signal sequences induces inversion of the intervening sequence. Small arrows indicate location of PCR primers to amplify joints [21].

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