Multiplexed cloning of long DNA sequences is a valuable technique in many biotechnology applications, such as long-read genome sequencing and the creation of open reading frame (ORF) libraries. Long-adapter single-stranded oligonucleotide (LASSO) probes have shown promise as a tool to clone long DNA fragments. LASSO probes are molecular inversion probes (MIP) engineered with an adapter region of user-defined length, flanked between template-specific probe sequences. Herein, we demonstrate that the adapter length is a key feature of LASSO that influences the efficiency of gene capture and cloning. Furthermore, we applied a model based on Monte Carlo molecular simulation in order to study the relationship between the long-adapter length of LASSO and capture enrichment. Our results suggest that the adapter length is a factor that contributes to the free energy of target-probe interaction, thereby determining the efficiency of capture. The results indicate that LASSOs with extremely long adapters cannot capture the targets well. They also suggest that targets of different lengths may prefer adapters of different lengths.
Keywords: DNA–DNA interaction; LASSO probe; VMMC simulation; gene-capture probe; oxDNA.