Targeted Sequencing of HPV16 in Cell Lines

For ligation sequencing 1 ug of cell line DNA was sheared to 8–20 kb with a G-tube (Covaris) or used unsheared with the LSK-109 kit. Targeted sequencing of HPV16 was carried out in CaSki and SiHa cells using CRISPR probes to HPV16 probes (Supplementary Table S1). For transposase sequencing of cell lines, 1 ug was used with the Rapid Sequencing kit (SQK-RAD004, Oxford Nanopore). CaSki and SNU-1000 DNA was also prepared using the Ultra-Long protocol of Circulomics and the Ultra-Long DNA Sequencing kit (SQK-ULK001, Oxford Nanopore) with and without adaptive sampling (35 (link),36 (link)) using a combined human HG38/high-risk HPV FASTA file selecting for cancer genes, integration loci, and HPV.
Tumor DNAs (0.3 ug) were sequenced using the Rapid Barcoding kit (SQK-RBK004, Oxford Nanopore). A total of 30–50 fM of DNA was loaded onto MinIon R9.4 flow cells (Oxford Nanopore).

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Lin M.H., Nguyen H.T., Dybala C, & Storti R.V. (1996). Myocyte-specific enhancer factor 2 acts cooperatively with a muscle activator region to regulate Drosophila tropomyosin gene muscle expression.. Proceedings of the National Academy of Sciences of the United States of America, 93(10), 4623-4628.

Publication 2023

Rapid Sequencing kit SQK-RAD004 Ultra-Long DNA Sequencing kit Rapid Barcoding kit SQK-RBK004 MinIon R9.4 flow cells

A dna A genes Adaptive Cancer Cell lines Cells Crispr Hpv16 Human Ligation Transposase Tumor dnas

Corresponding Organization :

Other organizations : National Cancer Institute, National Institutes of Health, Hospital General San Juan de Dios

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Variable analysis

independent variables

Shearing of cell line DNA to 8–20 kb with a G-tube (Covaris)
Use of unsheared cell line DNA with the LSK-109 kit
Use of 1 ug of cell line DNA for transposase sequencing with the Rapid Sequencing kit (SQK-RAD004, Oxford Nanopore)
Preparation of CaSki and SNU-1000 DNA using the Ultra-Long protocol of Circulomics and the Ultra-Long DNA Sequencing kit (SQK-ULK001, Oxford Nanopore) with and without adaptive sampling
Use of 0.3 ug of tumor DNA for sequencing using the Rapid Barcoding kit (SQK-RBK004, Oxford Nanopore)

dependent variables

Targeted sequencing of HPV16 in CaSki and SiHa cells using CRISPR probes to HPV16 probes
Sequencing of cell lines and tumor DNAs using various protocols and kits

control variables

Total of 30–50 fM of DNA loaded onto MinIon R9.4 flow cells (Oxford Nanopore)

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

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