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Protease and Proteinase Trials
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==4/9/10== ===Setup=== 2 columns/strips of sorted chromosomes were collected from Sam. The strips were separated as follows: | H | - Head Row 1 | o | - No chromosomes Row 2 | o | - 1 chromosome Row 3 | o | - 12 chromosomes Row 4 | o | - 12 chromosomes Row 5 | o | - 12 chromosomes Row 6 | o | - 23 chromosomes Row 7 | o | - 23 chromosomes Row 8 | o | - 23 chromosomes | A | - Tail The volumes of protease or proteinase K were the same in both strips: Rows 1 and 2: Nothing Row 3: 0.5uL P.K. (Positive Control) Row 4: 2uL P Row 5: 4uL P Row 6: 6uL P Row 7: 8uL P Row 8: Nothing (Negative Control) ===Incubation=== Strip 1: Heat denatured before adding proteases 95C, 10min -> 25C, 2min *Remove and add appropriate reagents (P. and P.K.) 37C, 23h36min Strip 2: No heat denaturing 37C, 23h36min ===Results=== '''Non-denatured samples:''' [[image: 12chr_PK.jpg]] ''Image 1: 12 chromosome, 0.5uL Proteinase K'' [[image: 23chr_8uL_P.jpg]] ''Image 2: 23 chromosome, 8uL Protease'' '''Denatured samples:''' [[image: preheat_12chr_PK.jpg]] ''Image 3: Denatured - 12 chromosome, 0.5uL Proteinase K'' [[image: preheat_23chr_6uL_P.jpg]] ''Image 4: Denatured - 23 chromosome, 6uL Protease'' [[image: preheat_23chr_8uL_P.jpg]] ''Image 5: Denatured - 23 chromosome, 8uL Protease'' ===Discussion=== While the results of this trial were somewhat less clear than previous trials, it appears that preheating and denaturing the samples before adding Protease aids in the denaturing capabilities of the Protease solution. On top of this, the difference between 6uL and 8uL of Protease is pretty small. To get higher amounts of Protease without overwhelming the volume of the chromosomes would be to reconstitute another vial with less water, increasing the concentration. This way we could test the effects of much higher amounts of protease without increasing the volume too much. Recommendation for future trials: Continue to preheat the samples before adding Protease, and use a more concentrated Protease solution.
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