Synthetic Biology for Plasmid Assembly
Artificial gene synthesis is a powerful tool, enabling modification of existing organisms with de novo assembled genetic elements to produce entirely novel expression products. When applied in an industrial setting, the possibilities for synthetic biology as a tool are vast. Building synthesized genes into plasmids allows for complete flexibility in downstream applications. Take this tool in house and have complete control over plasmid design and quality.
Challenge: Generate a plasmid pool with maximum diversity and possibilities in mind.
Artificially synthesized DNA constructs allow for custom gene design and optimization. Ordering oligonucleotides makes creating chimeric genes or codon-optimizing constructs for expression simple. In silico design of genes and plasmid components enables adding compatible sticky ends for Gibson or Golden Gate assembly and transposable construction of plasmids. Modular design of fragments greatly increases the possibilities for number of potential vectors, increasing the likelihood of downstream hits. Ligate combinations of fragments together to create a plasmid library.
Challenge: Create and select for bacterial producer strains carrying your plasmid.
Transform plasmids into a microbial plasmid production strain of choice. After allowing cells to recover in rich liquid media, plate onto an appropriate selection medium, targeting colonies carrying the plasmid with antibiotic resistance or a fluorescent marker. Pick colonies that successfully express the plasmid and passage into liquid culture to produce more plasmid for purification.
Challenge: Ensure your plasmid is a good candidate for further investment and ready for downstream applications.
Purify plasmids from liquid bacterial cultures to create a plasmid library of potential hits. Clean up plasmid DNA to remove bacterial endotoxins and residual salts and make plasmids transfection-grade. Create sequencing libraries of plasmids for NGS and perform initial expression analyses to determine the best candidates for investment and downstream use.
- Synthetic biology is a flexible and open-ended tool for producing new products.
- Gene fragments and plasmids must be modularly designed to maximize diversity of potential hits.
- Screen for candidate plasmids in a selection medium.
- Prepare plasmids for downstream application by cleaning up DNA for transfection and creating libraries for NGS. Confidently invest in the top candidate plasmids for further use. We can expedite and integrate touch points of this workflow together to ensure quick turnaround time to results.
Use the Biomek i-Series to automate complex steps of the protocol. Transform and culture bacterial cells and prepare DNA samples for use with this liquid handler. Perform initial testing of your product, from transduction or analytical characterization, and scale therapeutic biological production downstream of this workflow.
The Echo 525 Acoustic Liquid Handler flexes across every step of a synthetic biology workflow while miniaturizing reactions and consuming less resources. Array oligonucleotide fragments to construct plasmids, distribute bacterial cells across an agar plate for transformant selection, setup PCR reactions, create sequencing libraries of plasmids and test small molecules and antibodies.
Use the EMnetik 24 to accelerate and simplify nucleic acid isolation and cleanup with these microparticle processor or reagents. Paramagnetic bead technology selectively binds to DNA, securing it while contaminants like endotoxins and salts are washed away. Reduce error and variability inherent with protocols with manual pipetting.