OnRamp

OnRamp validates multiplexed plasmid sequences using Oxford Nanopore long-read sequencing for comprehensive detection of sequence variants and resolution of complex secondary structures.

Key Features:

• Long-read sequencing: Utilizes Oxford Nanopore technology to generate long reads enabling full-plasmid coverage and resolution of regions with high secondary structure.
• Multiplexing capability: Enables simultaneous sequencing and analysis of multiple plasmids in a single run.
• Full-plasmid coverage: Provides end-to-end plasmid sequence data suitable for validation of entire recombinant constructs.
• Alignment and quality metrics: Produces alignments between observed and predicted plasmid sequences and reports quality scores and read-level views.
• Scalability: Supports routine plasmid validation across multiple samples beyond single-plasmid assays.
• Sanger comparison: Addresses Sanger sequencing read-length and secondary-structure limitations by providing long-read alternatives for full-plasmid validation.

Scientific Applications:

• Plasmid validation: Confirms sequence integrity of recombinant plasmid vectors and detects sequence variations introduced during assembly.
• Molecular biology and genetic engineering: Applicable to construct verification in cloning, genome editing, and synthetic biology workflows.
• Proteomics and functional studies: Verifies coding sequences and regulatory elements for downstream protein expression and functional assays.
• High-throughput validation: Enables analysis of multiple plasmids or libraries using long-read sequencing data for routine verification.

Methodology:

A bioinformatics pipeline analyzes Oxford Nanopore long-read sequencing data to generate alignments to predicted plasmid sequences, compute quality scores, and produce read-level views.