NYMU Taipei/Lab Notes/2007 9 20
From 2007.igem.org
- (CinR+HSL D. ES #4) + D-term
- insert (CinR+HSL): 0.02 ug/uL = 20 ng/uL, 1530 bp
- vector (D-term): 0.06 ug/uL = 60 ng/uL, 3269 bp
- recommended vector mass: 100 ng to 150 ng
- MI/MV = C * SI/SV
- MI/MV = C * 1530/3269 =~ C/2
- assume VI = X, VV = Y, (X*20)/(Y*60) = C/2, X/Y = 3C/2
- total reaction volume is 20 uL and buffer is 10X
- for each reaction we need
- 1X buffer, thus volume of buffer is 20(uL)/10 = 2 (uL)
- 1 uL T4 DNA ligase (NEB, M0202)
- X + Y = 20 - (1+2) = 17 = (1 + 1.5C)Y
- choose C = 4, 7Y = 17, Y = 2.43, X = 6Y = 14.58
- approximately Y = 3, X = 14, C = 3.3
