Author: Long T. Nguyen; Brianna M Smith; Piyush K. Jain
Title: Enhancement of trans-cleavage activity of Cas12a with engineered crRNA enables amplified nucleic acid detection Document date: 2020_4_14
ID: n5kpvsvg_41
Snippet: The metal ions (Mg 2+ , Zn 2+ , Mn 2+ , Cu 2+ , Co 2+ , Ca 2+ ) were prepared by diluting chloride salt in different concentrations. For cis-cleavage, the Cas12a-crRNA-metal iron duplex was mixed with 100 nM: 100nM: varying nM ratio in 1X annealing buffer (100 mM Tris-HCl, pH 7.9 @ 25°C, 500 mM NaCl, 1 mg/ml BSA) and pre-incubated at 25 o C for 15 minutes. DNA activator (GFP or PCA3 fragments) was then added to the mixture to a total volume of 3.....
Document: The metal ions (Mg 2+ , Zn 2+ , Mn 2+ , Cu 2+ , Co 2+ , Ca 2+ ) were prepared by diluting chloride salt in different concentrations. For cis-cleavage, the Cas12a-crRNA-metal iron duplex was mixed with 100 nM: 100nM: varying nM ratio in 1X annealing buffer (100 mM Tris-HCl, pH 7.9 @ 25°C, 500 mM NaCl, 1 mg/ml BSA) and pre-incubated at 25 o C for 15 minutes. DNA activator (GFP or PCA3 fragments) was then added to the mixture to a total volume of 30 ïL and incubated at 37 o C for 30 minutes.
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