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10.1371/journal.pone.0190280

http://scihub22266oqcxt.onion/10.1371/journal.pone.0190280
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suck abstract from ncbi


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pmid29462136
      PLoS+One 2018 ; 13 (2 ): e0190280
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  • DNA methylation profiling of genomic DNA isolated from urine in diabetic chronic kidney disease: A pilot study #MMPMID29462136
  • Lecamwasam A ; Sexton-Oates A ; Carmody J ; Ekinci EI ; Dwyer KM ; Saffery R
  • PLoS One 2018[]; 13 (2 ): e0190280 PMID29462136 show ga
  • AIM: To characterise the genomic DNA (gDNA) yield from urine and quality of derived methylation data generated from the widely used Illuminia Infinium MethylationEPIC (HM850K) platform and compare this with buffy coat samples. BACKGROUND: DNA methylation is the most widely studied epigenetic mark and variations in DNA methylation profile have been implicated in diabetes which affects approximately 415 million people worldwide. METHODS: QIAamp Viral RNA Mini Kit and QIAamp DNA micro kit were used to extract DNA from frozen and fresh urine samples as well as increasing volumes of fresh urine. Matched buffy coats to the frozen urine were also obtained and DNA was extracted from the buffy coats using the QIAamp DNA Mini Kit. Genomic DNA of greater concentration than 20?g/ml were used for methylation analysis using the HM850K array. RESULTS: Irrespective of extraction technique or the use of fresh versus frozen urine samples, limited genomic DNA was obtained using a starting sample volume of 5ml (0-0.86?g/mL). In order to optimize the yield, we increased starting volumes to 50ml fresh urine, which yielded only 0-9.66?g/mL A different kit, QIAamp DNA Micro Kit, was trialled in six fresh urine samples and ten frozen urine samples with inadequate DNA yields from 0-17.7?g/mL and 0-1.6?g/mL respectively. Sufficient genomic DNA was obtained from only 4 of the initial 41 frozen urine samples (10%) for DNA methylation profiling. In comparison, all four buffy coat samples (100%) provided sufficient genomic DNA. CONCLUSION: High quality data can be obtained provided a sufficient yield of genomic DNA is isolated. Despite optimizing various extraction methodologies, the modest amount of genomic DNA derived from urine, may limit the generalisability of this approach for the identification of DNA methylation biomarkers of chronic diabetic kidney disease.
  • |*DNA Methylation [MESH]
  • |DNA/*urine [MESH]
  • |Diabetes Complications/*urine [MESH]
  • |Humans [MESH]
  • |Pilot Projects [MESH]


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