Project description:Here, we report an enrichment-based ultra-low input cfDNA methylation profiling method using methyl-CpG binding proteins capture, termed cfMBD-seq. We optimized the conditions of cfMBD capture by adjusting the amount of MethylCap protein along with using methylated filler DNA. Our data showed that cfMBD-seq performs equally to the standard MBD-seq (>1000 ng input) even when using 1 ng DNA as the input. cfMBD-seq demonstrated equivalent sequencing data quality as well as similar methylation profile when compared to cfMeDIP-seq. We showed that cfMBD-seq outperforms cfMeDIP-seq in the enrichment of CpG islands. This new bisulfite-free ultra-low input methylation profiling technology has a great potential in non-invasive and cost-effective cancer detection and classification.
Project description:Here we present Ultra-Mild Bisulfite Sequencing (UMBS-seq), which minimizes DNA damage and background noise while detecting 5-methylcytosine (5mC) in DNA. When applied to low-input lambda DNA (5 ng-10 pg), UMBS-seq achieved superior library yields, insert sizes, conversion efficiency, and CpG coverages compared to enzymatic alternatives. These results were recapitulated with low-input cell-free DNA (cfDNA) and in combination with targeted capture, highlighting the potential of UMBS-seq for future disease diagnosis.
Project description:In this study, the Infinium BeadChip DNA methylome profiling technology was optimized for suboptimal input DNA conditions, including ultra-low input down to single cells. We acquired knowledge on the boundaries and limits of the Infinium DNA methylation BeadChips with suboptimal input DNA. These findings offer comprehensive solutions to expand the application of this technology to cases with limited DNA.
2024-02-27 | GSE239290 | GEO
Project description:Short- and long-read sequencing of bacteriophages using ultra low starting DNA input
Project description:Ultra low input sequencing of FACS sorted primary murine microglia from CSF-1 or IL-34 deficient forebrain and cerebella, at P8 and 9 weeks
Project description:CD34+ hematopoietic stem progenitor cells (HSPCs) from cryo-preserved blood or bone marrow were FACS sorted in TriZol and RNA was isolated according to the manufacturer’s protocol. SMARTer Ultra Low Input RNA kit for sequencing (Clontech, v4 Cat# 634891) was used to generate cDNA. Sequencing libraries were generated using TruSeq Nano DNA Sample Preparation kits (Illumina, Cat# 20015964), according to the low sample protocol and paired-end sequenced on a HiSeq 2500 or Novaseq 6000 (both Illumina).
Project description:The STARR-seq plasmid library used for transfection was sequenced to enable input normalization. The sequencing library was generated with the NEBNext Ultra II FS DNA Library Prep Kit (New England Biolabs) and sequenced on the Illumina iSeq 100