POSoligo software for in vitro gene synthesis

• Published in: Scientific reports Vol. 14; no. 1; pp. 11117 - 8
• Main Authors: Tong, Yingying, Sun, Jie, Chen, Yang, Yi, Changhua, Wang, Hua, Li, Caixin, Dai, Nana, Yang, Guanghua
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 15.05.2024; Nature Publishing Group; Nature Portfolio
• Subjects: 631/1647/1511; 631/45/147; Algorithms; Bioinformatics; Computational Biology - methods; Conformation; COVID-19 - virology; Deoxyribonucleic acid; Design; DNA; DNA polymerase; DNA, Single-Stranded - chemistry; DNA, Single-Stranded - genetics; Experimental design; Humanities and Social Sciences; Kinases; Ligase chain reaction; multidisciplinary; Mutation rates; Nucleic acids; Nucleotide sequence; Nucleotide synthesis; Nucleotides; Oligonucleotides; Oligonucleotides - chemistry; Oligonucleotides - genetics; Patch oligonucleotide synthesis; Phosphorylation; Polymerase chain reaction; Polymerase Chain Reaction - methods; Protein structure; SARS-CoV-2 - genetics; Science; Science (multidisciplinary); Secondary structure; Severe acute respiratory syndrome coronavirus 2; Single-stranded DNA; Software; Software package; Nucleotide synthesis; Software package; Patch oligonucleotide synthesis
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-024-59497-3

Oligonucleotide synthesis is vital for molecular experiments. Bioinformatics has been employed to create various algorithmic tools for the in vitro synthesis of nucleotides. The main approach to synthesizing long-chain DNA molecules involves linking short-chain oligonucleotides through ligase chain reaction (LCR) and polymerase chain reaction (PCR). Short-chain DNA molecules have low mutation rates, while LCR requires complementary interfaces at both ends of the two nucleic acid molecules or may alter the conformation of the nucleotide chain, leading to termination of amplification. Therefore, molecular melting temperature, length, and specificity must be considered during experimental design. POSoligo is a specialized offline tool for nucleotide fragment synthesis. It optimizes the oligonucleotide length and specificity based on input single-stranded DNA, producing multiple contiguous long strands (COS) and short patch strands (POS) with complementary ends. This process ensures free 5′- and 3′-ends during oligonucleotide synthesis, preventing secondary structure formation and ensuring specific binding between COS and POS without relying on stabilizing the complementary strands based on Tm values. POSoligo was used to synthesize the linear RBD sequence of SARS-CoV-2 using only one DNA strand, several POSs for LCR ligation, and two pairs of primers for PCR amplification in a time- and cost-effective manner.