Dna nano 6000 assay kit

A total of 1.0 µg of DNA per sample was used as input material for library preparation. Genomic DNA was sonicated to a size of 350 bp, and then fragments were end-polished, A-tailed, and ligated with the full-length adapter for Illumina sequencing with further PCR amplification. At last, PCR products were purified (AMPure XP system) and libraries were analyzed for size distribution using the DNA Nano 6000 Assay Kit of Agilent Bioanalyzer 2100 system (Agilent Technologies, CA, USA) and quantified using real-time PCR. The Agilent SureSelect Human All Exome V6 was used to capture exome regions. A total of five samples, with low tumor cellularity (< 60%), were sequenced using a panel of 484 cancer-related genes (CGP; Cancer Gene Panel) generated by xGen™ Custom Hybridization Capture Panels (Integrated DNA Technologies, CA, USA). The DNA sequencing produced paired-end reads of 150 bp.

Genomic DNA was extracted and purified from whole blood using the Omega Bio-Tek E. Z.N.A. Blood DNA mini kit (Norcross, GA, United States). DNA concentration was measured using Qubit® DNA Assay Kit in Qubit® 2.0 Flurometer (Life Technologies, CA, United States). Fragment distribution of DNA library was measured using the DNA Nano 6000 Assay Kit of Agilent Bioanalyzer 2100 system (Agilent Technologies, CA, United States). A total amount of 1.0 μg DNA per sample was used as input material for the DNA sample preparations. Sequencing libraries were generated using NEBNext® DNA Library Prep Kit following manufacturer’s recommendations and indices were added to each sample. The genomic DNA is randomly fragmented to a size of 350 bp by shearing, then DNA fragments were end polished, A-tailed, and ligated with the NEBNext adapter for Illumina sequencing, and further PCR enriched by P5 and indexed P7 oligos. The PCR products were purified (AMPure XP system) and resultant libraries were analyzed for size distribution by Agilent 2100 Bioanalyzer and quantified using real-time PCR. Sequencing was performed on the Illumina platform (HiSeq X) using a paired-end read length of 150 bp. Data files have been uploaded to the European Nucleotide Archive with accession number PRJEB40922.

DNA degradation and contamination were monitored on 1% agarose gels. Then DNA purity was checked using the NanoPhotometer spectrophotometer (IMPLEN, Westlake Village, CA, United States). Subsequently, DNA concentration was measured using Qubit DNA Assay Kit in Qubit 2.0 Flurometer (Life Technologies, Carlsbad, CA, United States). Fragment distribution of DNA library was measured using the DNA Nano 6000 Assay Kit of Agilent Bioanalyzer 2,100 system (Agilent Technologies, Santa Clara, CA, United States).