Mimulus:Microsatellite recipes

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*The original reference for the AAT markers: [http://www.blackwell-synergy.com/doi/abs/10.1046/j.1365-294x.1998.00328.x Kelly and Willis 1998]  
*The original reference for the AAT markers: [http://www.blackwell-synergy.com/doi/abs/10.1046/j.1365-294x.1998.00328.x Kelly and Willis 1998]  
*Check out the [http://mimulusevolution.org Mimulus Biological Resource Center] for map location of these microsatellies.   
*Check out the [http://mimulusevolution.org Mimulus Biological Resource Center] for map location of these microsatellies.   
-
*The primers are archived in genbank.   
+
*The primers for these microsatellites are archived in genbank.
-
*Contact David Lowry (dbl2@duke.edu) for more inforamtion about these markers.
+
*See the [[Full Microsatellite Sequences]].
 +
*In addition to these recipes, check out the [[PCR Program for Microsatellites]].   
 +
*Contact David Lowry (dbl2@duke.edu) or other members of the [[Mimulus Community]] for more inforamtion about these markers.
 +
*[[Media:Microsatellite_PCR_conditions.xls|Microsat conditions spreadsheet]]
-
==Below are the recipes that have been fine-tuned for ''Mimulus guttatus'':==
+
Below are the recipes that have been fine-tuned for ''Mimulus guttatus''
==AAT 9==
==AAT 9==
Line 13: Line 16:
Cycles: 26
Cycles: 26
-
*H20 5.05μl
+
*H20 8.05μl
*10X Buffer 1.5μl
*10X Buffer 1.5μl
*MgCl2 (25mM) 1.2μl
*MgCl2 (25mM) 1.2μl
Line 19: Line 22:
*AAT9R 0.6μl
*AAT9R 0.6μl
*AAT9F 0.25μl
*AAT9F 0.25μl
-
*Taq 0.2μl
+
*Taq 0.2μl
-
 
+
==AAT 39==
==AAT 39==
Dye: FAM
Dye: FAM
-
 
Annealing Temp 53
Annealing Temp 53
Cycles 25
Cycles 25
-
 
*H20 5.05μl
*H20 5.05μl
Line 40: Line 40:
 +
==AAT 211==
 +
Dye: FAM
 +
Annealing Temp: 53
 +
Cycles: 22
 +
*H20 5.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*211R 0.6μl
 +
*211F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 217==
 +
Dye: FAM
 +
Annealing Temp: 54
 +
Cycles: 27
 +
*H20 8.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*217R 0.6μl
 +
*217F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 222==
 +
Dye: NED
 +
Annealing Temp 53
 +
Cycles: 30
 +
*H20 5.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*222R 0.6μl
 +
*222F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 225==
 +
Dye: VIC
 +
Annealing Temp: 53
 +
Cycles: 32
 +
*H20 8.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*225R 0.6μl
 +
*225F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 230==
 +
Dye: VIC
 +
Annealing Temp: 54
 +
Cycles: 28
 +
 +
*H20 6.85μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 2.4μl
 +
*dNTPs (.25mM) 1.2μl
 +
*230R 0.6μl
 +
*230F 0.25μl
 +
*Taq 0.2μl
-
AAT 211 - FAM
+
==AAT 233==
 +
Dye: FAM
 +
Annealing Temp: 53
 +
Cycles: 28
-
• Annealing Temp 53
+
*H20 5.05μl
-
• Cycles 22
+
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*233R 0.6μl
 +
*233F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 240==
 +
Dye: FAM
 +
Annealing Temp: 54
 +
Cycles: 25
-
    per sample x15 x110
+
*H20 7.45μl
-
H20 5.05μl 75.75μl 555.5μl
+
*10X Buffer 1.5μl
-
10X Buffer 1.5μl 22.5μl 165μl
+
*MgCl2 (25mM) 1.8μl
-
MgCl2 (25mM) 1.2μl 18μl 132μl
+
*dNTPs (.25mM) 1.2μl
-
dNTPs (.25mM) 1.2μl 18μl 132μl
+
*240R 0.6μl
-
• 211R 0.6μl 9μl 66μl
+
*240F 0.25μl
-
• 211F 0.25μl 3.75μl 27.5μl
+
*Taq 0.2μl
-
Taq 0.2μl 3μl 22μl
+
 +
==AAT 242==
 +
Dye: FAM
 +
Annealing Temp: 49
 +
Cycles: 32
 +
*H20 5.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*242R 0.6μl
 +
*242F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 261==
 +
Dye: VIC
 +
Annealing Temp: 49
 +
Cycles: 30
 +
*H20 4.45μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.8μl
 +
*dNTPs (.25mM) 1.2μl
 +
*261R 0.6μl
 +
*261F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 265==
 +
Dye: NED
 +
Annealing Temp: 55
 +
Cycles: 20
 +
*H20 5.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*265R 0.6μl
 +
*265F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 267==
 +
Dye: VIC
 +
Annealing Temp: 49
-
AAT 217 - FAM
+
Cycles: 30
 +
*H20 6.85μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 2.4μl
 +
*dNTPs (.25mM) 1.2μl
 +
*267R 0.6μl
 +
*267F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 272==
 +
Dye: NED
-
• Annealing Temp 54
 
-
• Cycles 27
 
 +
Annealing Temp: 54
 +
Cycles: 30
-
    per sample x15 x110
+
*H20 4.45μl
-
H20 5.05μl 75.75μl 555.5μl
+
*10X Buffer 1.5μl
-
10X Buffer 1.5μl 22.5μl 165μl
+
*MgCl2 (25mM) 1.8μl
-
MgCl2 (25mM) 1.2μl 18μl 132μl
+
*dNTPs (.25mM) 1.2μl
-
dNTPs (.25mM) 1.2μl 18μl 132μl
+
*272R 0.6μl
-
• 217R 0.6μl 9μl 66μl
+
*272F 0.25μl
-
• 217F 0.25μl 3.75μl 27.5μl
+
*Taq 0.2μl
-
Taq 0.2μl 3μl 22μl
+
 +
==AAT 278==
 +
Dye:  NED
 +
Annealing Temp: 53
 +
Cycles: 24
 +
 +
*H20 5.95μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.8μl
 +
*KCl (0.5M) 1.5μl
 +
*dNTPs (.25mM) 1.2μl
 +
*278R 0.6μl
 +
*278F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 283==
 +
Dye: VIC
 +
Annealing Temp: 52
 +
Cycles: 29
 +
*H20 3.85μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 2.4μl
 +
*dNTPs (.25mM) 1.2μl
 +
*283R 0.6μl
 +
*283F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 296==
 +
Dye: NED
 +
Annealing Temp: 53
 +
Cycles: 28
 +
*H20 2.1μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.8μl
 +
*KCl (.5M) 1.5μl
 +
*dNTPs (.25mM) 1.2μl
 +
*296R 1.2μl
 +
*296F 0.5μl
 +
*Taq 0.2μl
 +
==AAT 300==
 +
Dye: FAM
-
AAT 222- NED
+
Annealing Temp 55
 +
Cycles 22
-
• Annealing Temp 53
+
-
• Cycles 30
+
*H20 8.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*300R 0.6μl
 +
*300F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 308==
 +
Dye: VIC
 +
Annealing Temp: 55
-
    per sample x15 x110
+
Cycles: 24
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 222R 0.6μl 9μl 66μl
+
-
• 222F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
 +
*H20 5.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*308R 0.6μl
 +
*308F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 312==
 +
Dye: NED
 +
Annealing Temp: 51
 +
Cycles: 30
 +
 +
*H20 2.95μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.8μl
 +
*KCl (0.5M) 1.5μl
 +
*dNTPs (.25mM) 1.2μl
 +
*312R 0.6μl
 +
*312F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 333==
 +
Dye: VIC
 +
Annealing Temp: 51
-
AAT 225- VIC
+
Cycles: 30
 +
*H20 3.85μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 2.4μl
 +
*dNTPs (.25mM) 1.2μl
 +
*333R 0.6μl
 +
*Taq 0.2μl
-
• Annealing Temp 53
 
-
• Cycles 32
 
 +
==AAT 350==
 +
Dye: FAM
 +
Annealing Temp: 52
-
    per sample x15 x110
+
Cycles: 28
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 225R 0.6μl 9μl 66μl
+
-
• 225F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
 +
*H20 5.05μl
 +
*10X Buffer 1.5μl
 +
*MgCl2 (25mM) 1.2μl
 +
*dNTPs (.25mM) 1.2μl
 +
*350R 0.6μl
 +
*350F 0.25μl
 +
*Taq 0.2μl
 +
==AAT 356==
 +
Dye: VIC
 +
Annealing Temp: 55
-
 
+
Cycles: 25
-
 
+
-
 
+
*H20 8.05μl
-
 
+
*10X Buffer 1.5μl
-
 
+
*MgCl2 (25mM) 1.2μl
-
 
+
*dNTPs (.25mM) 1.2μl
-
 
+
*356R 0.6μl
-
 
+
*356F 0.25μl
-
 
+
*Taq 0.2μl
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 230 - VIC
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 54
+
-
• Cycles 28
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 3.85μl 57.75μl 423.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 2.4μl 36μl 264μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 230R 0.6μl 9μl 66μl
+
-
• 230F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 233- FAM
+
-
 
+
-
 
+
-
• Annealing Temp 53
+
-
• Cycles 28
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 233R 0.6μl 9μl 66μl
+
-
• 233F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 240 - FAM
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 54
+
-
Cycles 25
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
H20 4.45μl 66.75μl 489.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.8μl 27μl 198μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 240R 0.6μl 9μl 66μl
+
-
• 240F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 242- FAM
+
-
 
+
-
 
+
-
• Annealing Temp 49
+
-
• Cycles 32
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 5.05μl 75.75μl 555.5μl
+
-
10X Buffer 1.5μl 22.5μl 165μl
+
-
MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 242R 0.6μl 9μl 66μl
+
-
• 242F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 261 - VIC
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 49
+
-
• Cycles 30
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 4.45μl 66.75μl 489.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.8μl 27μl 198μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 261R 0.6μl 9μl 66μl
+
-
• 261F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 265- NED
+
-
 
+
-
 
+
-
• Annealing Temp 55
+
-
• Cycles 20
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 265R 0.6μl 9μl 66μl
+
-
• 265F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 267- VIC
+
-
 
+
-
 
+
-
• Annealing Temp 49
+
-
• Cycles 30
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 3.85μl 57.75μl 423.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 2.4μl 36μl 264μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 267R 0.6μl 9μl 66μl
+
-
• 267F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 272- NED
+
-
 
+
-
 
+
-
• Annealing Temp 54
+
-
• Cycles 30
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 4.45μl 66.75μl 489.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.8μl 27μl 198μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 272R 0.6μl 9μl 66μl
+
-
• 272F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 278 - NED
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 53
+
-
• Cycles 24
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 2.95μl 44.25μl 324.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.8μl 27μl 198μl
+
-
• KCl (0.5M) 1.5μl 22.5μl 165μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 278R 0.6μl 9μl 66μl
+
-
• 278F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 283 - VIC
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 52
+
-
• Cycles 29
+
-
 
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 3.85μl 96.25μl 423.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 2.4μl 36μl 264μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 283R 0.6μl 9μl 66μl
+
-
• 283F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 296 - NED
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 53
+
-
• Cycles 28
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 2.1μl 31.5μl 231μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.8μl 27μl 198μl
+
-
• KCl (.5M) 1.5μl 22.5μl 165μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 296R ** 1.2μl 18μl 132μl **
+
-
• 296F ** 0.5μl 7.5μl 55μl **
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 300 - FAM
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 55
+
-
• Cycles 22
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 300R 0.6μl 9μl 66μl
+
-
• 300F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 308 - VIC
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 55
+
-
• Cycles 24
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 308R 0.6μl 9μl 66μl
+
-
• 308F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 312 - NED
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 51
+
-
• Cycles 30
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 2.95μl 44.25μl 324.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.8μl 27μl 198μl
+
-
• KCl (0.5M) 1.5μl 22.5μl 165μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 312R 0.6μl 9μl 66μl
+
-
• 312F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 333 - VIC
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 51
+
-
• Cycles 30
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 3.85μl 57.75μl 423.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 2.4μl 36μl 264μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 333R 0.6μl 9μl 66μl
+
-
• 333F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 350 - FAM
+
-
 
+
-
 
+
-
 
+
-
• Annealing Temp 52
+
-
• Cycles 28
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
• 350R 0.6μl 9μl 66μl
+
-
• 350F 0.25μl 3.75μl 27.5μl
+
-
• Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
 
+
-
AAT 356- VIC
+
-
 
+
-
 
+
-
• Annealing Temp 55
+
-
• Cycles 25
+
-
 
+
-
 
+
-
 
+
-
 
+
-
    per sample x15 x110
+
-
• H20 5.05μl 75.75μl 555.5μl
+
-
• 10X Buffer 1.5μl 22.5μl 165μl
+
-
• MgCl2 (25mM) 1.2μl 18μl 132μl
+
-
• dNTPs (.25mM) 1.2μl 18μl 132μl
+
-
356R 0.6μl 9μl 66μl
+
-
356F 0.25μl 3.75μl 27.5μl
+
-
Taq 0.2μl 3μl 22μl
+
-
 
+
-
 
+
-
 
+
-
 
+

Current revision

Below are the recipes that have been fine-tuned for Mimulus guttatus

Contents

AAT 9

Dye: VIC

Annealing Temp: 55

Cycles: 26

  • H20 8.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (2.5mM) 1.2μl
  • AAT9R 0.6μl
  • AAT9F 0.25μl
  • Taq 0.2μl

AAT 39

Dye: FAM

Annealing Temp 53

Cycles 25

  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 39R 0.6μl
  • 39F 0.25μl
  • Taq 0.2μl


AAT 211

Dye: FAM

Annealing Temp: 53

Cycles: 22

  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 211R 0.6μl
  • 211F 0.25μl
  • Taq 0.2μl


AAT 217

Dye: FAM

Annealing Temp: 54

Cycles: 27


  • H20 8.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 217R 0.6μl
  • 217F 0.25μl
  • Taq 0.2μl

AAT 222

Dye: NED

Annealing Temp 53

Cycles: 30

  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 222R 0.6μl
  • 222F 0.25μl
  • Taq 0.2μl

AAT 225

Dye: VIC

Annealing Temp: 53

Cycles: 32

  • H20 8.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 225R 0.6μl
  • 225F 0.25μl
  • Taq 0.2μl

AAT 230

Dye: VIC

Annealing Temp: 54

Cycles: 28


  • H20 6.85μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 2.4μl
  • dNTPs (.25mM) 1.2μl
  • 230R 0.6μl
  • 230F 0.25μl
  • Taq 0.2μl

AAT 233

Dye: FAM

Annealing Temp: 53

Cycles: 28

  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 233R 0.6μl
  • 233F 0.25μl
  • Taq 0.2μl

AAT 240

Dye: FAM

Annealing Temp: 54

Cycles: 25

  • H20 7.45μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.8μl
  • dNTPs (.25mM) 1.2μl
  • 240R 0.6μl
  • 240F 0.25μl
  • Taq 0.2μl

AAT 242

Dye: FAM

Annealing Temp: 49

Cycles: 32


  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 242R 0.6μl
  • 242F 0.25μl
  • Taq 0.2μl


AAT 261

Dye: VIC


Annealing Temp: 49

Cycles: 30


  • H20 4.45μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.8μl
  • dNTPs (.25mM) 1.2μl
  • 261R 0.6μl
  • 261F 0.25μl
  • Taq 0.2μl


AAT 265

Dye: NED


Annealing Temp: 55

Cycles: 20

  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 265R 0.6μl
  • 265F 0.25μl
  • Taq 0.2μl


AAT 267

Dye: VIC


Annealing Temp: 49

Cycles: 30

  • H20 6.85μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 2.4μl
  • dNTPs (.25mM) 1.2μl
  • 267R 0.6μl
  • 267F 0.25μl
  • Taq 0.2μl

AAT 272

Dye: NED


Annealing Temp: 54

Cycles: 30

  • H20 4.45μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.8μl
  • dNTPs (.25mM) 1.2μl
  • 272R 0.6μl
  • 272F 0.25μl
  • Taq 0.2μl


AAT 278

Dye: NED

Annealing Temp: 53

Cycles: 24



  • H20 5.95μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.8μl
  • KCl (0.5M) 1.5μl
  • dNTPs (.25mM) 1.2μl
  • 278R 0.6μl
  • 278F 0.25μl
  • Taq 0.2μl

AAT 283

Dye: VIC

Annealing Temp: 52

Cycles: 29

  • H20 3.85μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 2.4μl
  • dNTPs (.25mM) 1.2μl
  • 283R 0.6μl
  • 283F 0.25μl
  • Taq 0.2μl


AAT 296

Dye: NED

Annealing Temp: 53

Cycles: 28


  • H20 2.1μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.8μl
  • KCl (.5M) 1.5μl
  • dNTPs (.25mM) 1.2μl
  • 296R 1.2μl
  • 296F 0.5μl
  • Taq 0.2μl


AAT 300

Dye: FAM

Annealing Temp 55

Cycles 22


  • H20 8.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 300R 0.6μl
  • 300F 0.25μl
  • Taq 0.2μl

AAT 308

Dye: VIC

Annealing Temp: 55

Cycles: 24

  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 308R 0.6μl
  • 308F 0.25μl
  • Taq 0.2μl


AAT 312

Dye: NED

Annealing Temp: 51

Cycles: 30

  • H20 2.95μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.8μl
  • KCl (0.5M) 1.5μl
  • dNTPs (.25mM) 1.2μl
  • 312R 0.6μl
  • 312F 0.25μl
  • Taq 0.2μl


AAT 333

Dye: VIC

Annealing Temp: 51

Cycles: 30

  • H20 3.85μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 2.4μl
  • dNTPs (.25mM) 1.2μl
  • 333R 0.6μl
  • Taq 0.2μl


AAT 350

Dye: FAM

Annealing Temp: 52

Cycles: 28

  • H20 5.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 350R 0.6μl
  • 350F 0.25μl
  • Taq 0.2μl


AAT 356

Dye: VIC

Annealing Temp: 55

Cycles: 25

  • H20 8.05μl
  • 10X Buffer 1.5μl
  • MgCl2 (25mM) 1.2μl
  • dNTPs (.25mM) 1.2μl
  • 356R 0.6μl
  • 356F 0.25μl
  • Taq 0.2μl





AAT 364- NED


• Annealing Temp 53 • Cycles 30



    per sample	x15		x110	

• H20 3.7μl 55.5μl 407μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 1.8μl 27μl 198μl • KCl (.5M) 0.75μl 11.25μl 82.5μl • dNTPs (.25mM) 1.2μl 18μl 132μl • 364R 0.6μl 9μl 66μl • 364F 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl













AAT 367 - VIC


• Annealing Temp 54 • Cycles 22


    per sample	x15		x110	

• H20 4.45μl 66.75μl 489.5μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 1.8μl 27μl 198μl • dNTPs (.25mM) 1.2μl 18μl 132μl • 367R 0.6μl 9μl 66μl • 367F 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl











AAT 372- NED


• Annealing Temp 55 • Cycles 25



    per sample	x15		x110	

• H20 4.45μl 66.75μl 621.5μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 0.6μl 9μl 66μl • dNTPs (.25mM) 1.2μl 18μl 132μl • 372R 0.6μl 9μl 66μl • 372F 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl










AAT 374 - FAM


• Annealing Temp 53 • Cycles 28


    per sample	x15		x110	

• H20 4.45μl 66.75μl 489.5μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 1.8μl 27μl 198μl • dNTPs (.25mM) 1.2μl 18μl 132μl • 374R 0.6μl 9μl 66μl • 374F 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl











AG 19- NED


• Annealing Temp 53 • Cycles 30



    per sample	x15		x110	

• H20 5.65μl 84.75μl 621.5μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 0.6μl 9μl 66μl • dNTPs (.25mM) 1.2μl 18μl 132μl • AG19R 0.6μl 9μl 66μl • AG19F 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl










ap3-VIC


• Annealing Temp 53 • Cycles 25


    per sample	x15		x110	

• H20 5.1μl 76.5μl 561μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 1.2μl 18μl 132μl • dNTPs (.25mM) 1.2μl 18μl 132μl • AP3R 0.6μl 9μl 66μl • AP3F 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl










lfy- NED


• Annealing Temp 52 • Cycles 19


    per sample	x15		x110	

• H20 5.05μl 75.75μl 555.5μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 1.2μl 18μl 132μl • dNTPs (.25mM) 1.2μl 18μl 132μl • LFYR 0.6μl 9μl 66μl • LFYF 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl







    CYCB - FAM


• Annealing temp 62 • cycles 16


    per sample	x15		x110	

• H20 5.05μl 75.75μl 555.5μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 1.2μl 18μl 132μl • dNTPs (.25mM) 1.2μl 18μl 132μl • CYCBR 0.6μl 9μl 66μl • CYCBF 0.25μl 3.75μl 27.5μl • Taq 0.2μl 3μl 22μl








    CYCA - FAM


• Annealing temp 58 • cycles 37


    per sample	x15		x110	

• H20 4.05μl 60.75μl 445.5μl • 10X Buffer 1.5μl 22.5μl 165μl • MgCl2 (25mM) 1.8μl 27μl 198μl • dNTPs (.25mM) 1.5μl 22.5μl 165μl • CYCAR 0.65μl 9.75μl 71.5μl • CYCAF 0.3μl 4.5μl 33μl • Taq 0.2μl 3μl 22μl

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