User:Cesar A. Villavicencio/Notebook/Tyrosinase Extraction/2012/05/01
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Primer Order and Design6 Primers were designed for tyrosinase extraction from Bacillus megaterium QMB1551 for the extraction and in situ mutation of the gene. The primers designed were the following Primers for Tyrosinase Extraction TyrFWD 5'-GAATTCGCCCCTTGCTTCTTT-3' Length=21 A=2 T=9 C=7 G=3 CG=47.62% Linguistic Complexity=82% Primer's PCR Efficiency=85% Tm=56.4°C (Allawi's thermodynamics parameters)** Tm=56.4°C (Tm = 77.1 + 11.7Log[K+] + 0.41(GC%) - 528/L) Extinction coefficient = 178200 L/(mol•cm) Molecular weight = 6314.13 g/mol
5'-CTGCTTTTGATTGCATACCG-3' Length=20 A=3 T=8 C=5 G=4 CG=45% Linguistic Complexity=85% Primer's PCR Efficiency=85% Tm=52.9°C (Allawi's thermodynamics parameters)** Tm=54.3°C (Tm = 77.1 + 11.7Log[K+] + 0.41(GC%) - 528/L) Extinction coefficient = 179100 L/(mol•cm) Molecular weight = 6073.98 g/mol
Primers for in situ mutation MutTyrFWD 5'-gacgcacagctccaagatccctcacag-3' Length=27 A=8.0 G=5.0 T=3.0 C=11.0 CG=59.3% Linguistic complexity = 78% Primer's PCR efficiency = 88%
Tm = 69.1°C (Sugimoto's thermodynamics parameters) Tm = 52.1°C (SantaLucia's thermodynamics parameters) Tm = 64.6°C (Tm = 75.1 + 11.7Log[K+] + 0.41(GC%) - 528/L) Tm = 64.3°C (Tm = 64.9 + 41(nG + nC - 16.4)/L)
MutTyrREV 5'-ctgtgagggatcttggagctgtgcgtc-3' Length=27 A=3.0 G=11.0 T=8.0 C=5.0 CG=59.3% Linguistic complexity = 78% Primer's PCR efficiency = 93% Tm = 64.1°C (Allawi's thermodynamics parameters) Tm = 69.1°C (Sugimoto's thermodynamics parameters) Tm = 57.6°C (SantaLucia's thermodynamics parameters) Tm = 64.6°C (Tm = 75.1 + 11.7Log[K+] + 0.41(GC%) - 528/L) Tm = 64.3°C (Tm = 64.9 + 41(nG + nC - 16.4)/L)
Primers for BioBrick adaptation BMTyrFWD 5'-gaattcgcggccgcttctagaggaaagaggttaaaaaatgagtaacaagtac-3' Length=52 A=20.0 G=14.0 T=10.0 C=8.0 CG=42.3% Linguistic complexity = 82% Primer's PCR efficiency = 39% Tm = 65.9°C (Allawi's thermodynamics parameters) Tm = 73.4°C (Sugimoto's thermodynamics parameters) Tm = 62.2°C (SantaLucia's thermodynamics parameters) Tm = 67.1°C (Tm = 75.1 + 11.7Log[K+] + 0.41(GC%) - 528/L) Tm = 69.3°C (Tm = 64.9 + 41(nG + nC - 16.4)/L)
BMTyrREV 5'-ctgcagcggccgctactagtaaacgggttatgatgaacgtt-3' Length=41 A=10.0 G=12.0 T=10.0 C=9.0 CG=51.2% Linguistic complexity = 86% Primer's PCR efficiency = 17%
Tm = 74.4°C (Sugimoto's thermodynamics parameters) Tm = 62.9°C (SantaLucia's thermodynamics parameters) Tm = 68.0°C (Tm = 75.1 + 11.7Log[K+] + 0.41(GC%) - 528/L) Tm = 69.5°C (Tm = 64.9 + 41(nG + nC - 16.4)/L)
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