3/11/2023 0 Comments Integrity plus pool repair![]() ![]() Here we report the success of this approach and describe the resulting phenotype with respect to responses to a variety of DNA-damaging agents. For these reasons we sought to develop a cell line in which the UVSSA gene was inactivated by CRISPR-Cas9 targeting of the first coding exon. An additional complication is that some mutations in XP and CS genes also give rise to the UV s clinical phenotype ( 13, 14). This is because of the rarity of available fibroblast cell lines and the difficulty of recognizing the subtle clinical phenotype. Although there has been extensive analysis of the phenotypes of mutations in CSA and CSB in response to transcription-blocking lesions and oxidative damage, less is known about the phenotype of mutations in UVSSA. TCR activates a cycle of ubiquitinylation and deubiquitinylation of CSB by the CSA ligase complex followed by recruitment of the USP7 protease by UVSSA ( 12). TCR involves at least four core proteins: CSA (ERCC8), CSB (ERCC6), UVSSA (KIAA1530), and USP7. We anticipate that this knockout cell line will advance understanding of this and possibly related transcription-coupled DNA repair diseases. We discuss this in terms of the likely commutative interplay of factors in CS. Loss of UVSSA protein did not, however, increase sensitivity to oxidative damage or to inhibitors of poly (ADP)ribose polymerase, unlike reported in CSB cells. Transcription arrest in a UVSSA knockout cell line resulted in ATM-dependent phosphorylation of H2Ax and delayed DNA synthesis, relieved by an inhibitor of ATM. Elimination of the UVSSA protein was confirmed by Western blotting and the knockout cells displayed the predicted sensitivity to transcription blocking lesions caused by illudin, cisplatin, and ultraviolet light, just as in CS cell lines. In this study we deployed CRISPR to disrupt exon I of the UVSSA gene in the human embryonic kidney cell line HEK293. Mutations in UVSSA, in contrast, are found in the rare mild photosensitive syndrome (UV s) that lacks the noncutaneous complications of CSA or CSB patients. Mutations in CSA and CSB are found in Cockayne syndrome (CS), which is a human recessively inherited photosensitive, neurocutaneous, aging disorder. Very satisfied.The UVSSA (KIAA1530) protein is a component of transcription-coupled repair which, together with the CSA(ERCC8) and CSB(ERCC6) proteins cooperates to relieve transcription-blocking DNA damage. ![]() Please see some of my before and after pictures to get an idea of the scope of the work. ![]() ![]() All in all, I'm very happy with the way the project came out and I would definitely do business with West Coast Pools again for any work I need done in the future. Also, throughout the entire project David was there to answer any questions that I had to ensure I was happy with the way the project was coming along. There was actually one area of the job I wasn't completely happy with but David had a member of his team come out and complete the job to my standards. David's team came out and did a great job and completed the project within the specified time frame. What I especially like about David is that he doesn't try to up-sell you on services that you don't need, instead, he focuses solely on what your needs are based on the scope of the project and within the confines of your budget. David arrived at my house in a timely manner and provided me a fair and accurate estimate of the project cost as well as an estimated start date. cleaning, lighting replacement and some stucco work. My pool needed close to 10k worth of work including coping repair, mastic replacement, complete stone replacement, tile. ![]()
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