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A detailed purification strategy for thaumatin is reported resulting in a homogenous sample recovered at a yield of 42%.
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Detailed protein purifications, antibodies used and procedures for electron microscopy and single-particle analysis are included in the Supplementary Materials and Methods.
We present detailed protocols for the purification of all essential protein components of the minimal eukaryotic transcription initiation complex.
The lysates were then subjected to purification as detailed in Materials and Methods.
SMRTbell libraries were constructed using end-repair, ligation, and exonuclease purification strategies detailed in the Pacific Biosciences P5-C3 TemPreparationration Kit protocols.
Both proteins were expressed in Escherichia coli as N-terminal glutathione- S-transferase (GST -fusion proteins before cleavaGST -fusionfication as detailed under Materials and methods.
Additional tables and figures, detailed methods for protein purification, assay conditions (including a table summarizing all of the reaction conditions used for dose response), methods for isothermal titration calorimetry and cellular assays.
We have found that this method of purification greatly reduces DNA loss (~8.6% loss after each purification step) as compared to column purification (~20.5% loss after each purification step) and standard SPRI purification (~18.8% loss after each purification step), as detailed in Additional file 1.
This is the first report detailing the successful purification of a genetically engineered earthworm fibrinolytic enzyme.
Herein, we provide detailed protocols for engineering and purification of a cell-permeant FLP recombinase protein.
The whole purification procedure is detailed in Figure 1.
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