title: Merlin: Computer-Aided Oligonucleotide Design for Large Scale Genome Engineering with MAGE.
abstract: Genome engineering technologies now enable precise manipulation of organism genotype, but can be limited in scalability by their design requirements. Here we describe Merlin ( http://merlincad.org ), an open-source web-based tool to assist biologists in designing experiments using multiplex automated genome engineering (MAGE). Merlin provides methods to generate pools of single-stranded DNA oligonucleotides (oligos) for MAGE experiments by performing free energy calculation and BLAST scoring on a sliding window spanning the targeted site. These oligos are designed not only to improve recombination efficiency, but also to minimize off-target interactions. The application further assists experiment planning by reporting predicted allelic replacement rates after multiple MAGE cycles, and enables rapid result validation by generating primer sequences for multiplexed allele-specific colony PCR. Here we describe the Merlin oligo and primer design procedures and validate their functionality compared to OptMAGE by eliminating seven AvrII restriction sites from the Escherichia coli genome.
authors.affiliation: Program in Bioinformatics, Boston University , Boston, Massachusetts 02215, United States.|Biological Design Center (BDC), Boston University , Boston, Massachusetts 02215, United States.
authors.forename: Michael
authors.initials: M
authors.lastname: Quintin
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, Yale University , New Haven 06511, Connecticut, United States.|Systems Biology Institute, Yale University West Campus , West Haven, Connecticut 06516, United States.
authors.forename: Natalie J
authors.initials: NJ
authors.lastname: Ma
authors.affiliation: Department of Electrical and Computer Engineering, Boston University , Boston, Massachusetts 02215, United States.
authors.forename: Samir
authors.initials: S
authors.lastname: Ahmed
authors.affiliation: Department of Electrical and Computer Engineering, Boston University , Boston, Massachusetts 02215, United States.
authors.forename: Swapnil
authors.initials: S
authors.lastname: Bhatia
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, Yale University , New Haven 06511, Connecticut, United States.
authors.forename: Aaron
authors.initials: A
authors.lastname: Lewis
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, Yale University , New Haven 06511, Connecticut, United States.|Systems Biology Institute, Yale University West Campus , West Haven, Connecticut 06516, United States.
authors.forename: Farren J
authors.initials: FJ
authors.lastname: Isaacs
authors.affiliation: Department of Electrical and Computer Engineering, Boston University , Boston, Massachusetts 02215, United States.
authors.forename: Douglas
authors.initials: D
authors.lastname: Densmore
mesh_terms: D017076:Computer-Aided Design; D017931:DNA Primers / Q000235:genetics; D004277:DNA, Single-Stranded / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D016680:Genome, Bacterial / Q000235:genetics; D023281:Genomics / Q000379:methods*; D020407:Internet; D009841:Oligonucleotides / Q000235:genetics*; D011995:Recombination, Genetic / Q000235:genetics; D012107:Research Design; D012984:Software
chemical_list: D017931:DNA Primers; D004277:DNA, Single-Stranded; D009841:Oligonucleotides
keywords: MAGE; automation; bioengineering; genomics; multiplex automated genome engineering; software; synthetic biology