Objective

Utilizing the Labcyte Echo 525 liquid handler we demonstrate an efficient and robust gene assembly method to generate a fusion protein for functional cellular assays. A PTEN-GFP construct was generated using the Gateway chemistry. PTEN null PC-3 cells were transfected with this construct to generate a fluorescently-labelled PTEN cell line to interrogate the PI3K pathway. The DNA from these cells was purified and the construct integration into the genome was confirmed by whole genome sequencing. The assembled fusion protein now enables functional cellular assays including, but not limited to, viability and localization studies. The Echo 525 liquid handler enables flexibility and scalability for gene assembly while streamlining and automating the process. The library preparation was miniaturized and streamlined for automation by using the Echo System’s acoustic, non-contact dispensing eliminates tips and cross-contamination. The low 25 nL resolution enables reduction in reaction volume which results in reduced reagent consumption and reduced costs for the user.