As there is halo zone already appeared around the colony which was incubated at 370C on tributyrin plate. The colony of native lipase shows halo zone at 370C too. Please suggest a mutant screening method to identify thermostable lipase.
* Native lipase was subjected to error prone PCR ,randomly introduce mutations to enhance thermostability of the gene.
Stab isolated colonies with a toothpick. Add toothpick to media in tubes and incubate for 2hr at 50oC. Add a drop of the media to tributyrin plates.
you can try my software.
A Method to Rationally Increase Protein Stability Based on the Charge–Charge Interaction, with Application to Lipase LipK107. Protein Science, 2014, 23(1):110-116.
For lipase assay I would like to advise to to follow winkler and stuckman 1979 methos as given in these papers.
https://www.researchgate.net/publication/47644372_Synthesis_of_ethyl_ferulate_in_organic_medium_using_Celite-immobilized_lipase
https://www.researchgate.net/publication/224900014_Catalytic_Potential_of_a_Nitrocellulose_Membrane-_Immobilized_Lipase_in_Aqueous_and_Organic_Media?ev=prf_pub
https://www.researchgate.net/publication/284078828_Cellulose_binding_domain_assisted_immobilization_of_lipase_GSlip-CBD_onto_cellulosic_nanogel_Characterization_and_application_in_organic_medium?ev=prf_pub
Could I get some advice on Lipase Assay? - ResearchGate. Available from: https://www.researchgate.net/post/Could_I_get_some_advice_on_Lipase_Assay#58a2c1b7dc332d157d7afa63 [accessed Feb 14, 2017].
Article Synthesis of ethyl ferulate in organic medium using Celite-i...
Article Catalytic Potential of a Nitrocellulose Membrane- Immobilize...
Article Cellulose binding domain assisted immobilization of lipase (...
To screen the lipase you can use ).5% tributyrin agar plate.
Thanks
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