Leap-In transposases integrate single copies of the entire synthetic transposon into multiple transcriptionally active genomic loci. Expression constructs integrated by Leap-In transposases do not exhibit the rearrangements and concatemerization that are common, problematic hallmarks of random non-homologous recombination driven integrations. Maintaining structural integrity of the sequence integrated into the host genome ensures that regulatory elements remain associated with the appropriate open reading frames, and desired balances between multiple open reading frames are maintained.
The balance of expression for each subunit in a multi-subunit protein affects productivity and product quality. Compounding this complexity, there is often an incomplete correlation between the best ratios of transiently transfected genes, and the optimal stable expression of each subunit. Using our VectorGPS® design platform, we can design, synthesize and transfect several synthetic transposons in parallel, each harboring up to 4 independently-controlled, codon-optimized ORFs with altered expression ratios. Stable pool productivity and product quality analyses will reveal the optimal construct.
Control of protein subunit expression ratios in 2-, 3- and 4-ORF Leap-In transposons.
ATUM currently uses two host cell lines with lineage traceability and documented provenance, are adapted to commercially-available chemically-defined serum-free formulations. No raw materials of animal origin are used during our CLD process.
Leap-In transposase is very efficient. Most cells that are transfected with Leap-In transposons and transposase will integrate multiple copies of the transposon into their genomes. This results in rapid recovery of stable pools consisting of many very similar cells. This means that the productivity and product quality of clones derived from these pools are similar to the pools themselves. Process and analytical development can therefore start as early as a few weeks post-transfection. Material for toxicology studies can be prepared from stable pools to further shorten overall CMC timelines.
Product quality (glycan relative abundance %, charge %) and productivity is comparable between stable pool and derivative clones.
In stable pools generated by Leap-In technology, the clonal distributions within the pools are strongly biased towards high productivity. This reduces the number of clones that need to be tested to at most a few hundred.
Static 96 well plate
24 deep-well scale, 7 day fed batch clone ranking
Highest ranked clones, 125 ml shake, 14 day fed batch
Screening, characterizing and ranking fewer clones eliminates the need to invest in expensive, high-throughput instrumentation and allows more cell line development projects to be managed simultaneously.