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Thermostable ReBphP-PCM (2022_Q3-4) - Coggle Diagram
Thermostable ReBphP-PCM (2022_Q3-4)
Random Mutagenesis
Error-Prone PCR optimization
Error-prone PCR + Gibson ligation
High-throughput plate screening (SWIR setup)
Follow-up validation of selected clones
Re-streaking + Re-testing in SWIR setup
Mini-prep + sequencing
Expression on plate
37°C
SDS-PAGE (expression levels)
Expression in liq. culture
Batch Purification
Photophysics
Cleared Lysate
Targeted Mutagenesis
Literature review of BphPs for structural and sequence indications on thermostability
Design + Cloning of RePCM clones with mutations of interest
Expression on plate
37°C
SDS-PAGE (expression levels)
Expression in liq. culture
Batch Purification
Photophysics
Cleared Lysate
Domain Expression dissection
Comparative Stability tests:
RePCM (P540) and DrPCM (P685)
Mammalian Expression & Stability
Dual Expression Vectors