Difference between revisions of "Mimulus:Microsatellite recipes"

From OpenWetWare
Jump to: navigation, search
(AAT 267)
 
(13 intermediate revisions by 2 users not shown)
Line 5: Line 5:
 
*In addition to these recipes, check out the [[PCR Program for Microsatellites]].   
 
*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.
 
*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 15: 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 21: Line 22:
 
*AAT9R 0.6μl
 
*AAT9R 0.6μl
 
*AAT9F 0.25μl
 
*AAT9F 0.25μl
*Taq 0.2μl
+
*Taq 0.2μl
 
 
  
 
==AAT 39==
 
==AAT 39==
Line 63: Line 63:
  
  
*H20 5.05μl
+
*H20 8.05μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 1.2μl
 
*MgCl2 (25mM) 1.2μl
Line 69: Line 69:
 
*217R 0.6μl
 
*217R 0.6μl
 
*217F 0.25μl
 
*217F 0.25μl
*Taq 0.2μl
+
*Taq 0.2μl
 
 
  
 
==AAT 222==
 
==AAT 222==
Line 93: Line 92:
 
Cycles: 32
 
Cycles: 32
  
*H20 5.05μl
+
*H20 8.05μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 1.2μl
 
*MgCl2 (25mM) 1.2μl
Line 99: Line 98:
 
*225R 0.6μl
 
*225R 0.6μl
 
*225F 0.25μl
 
*225F 0.25μl
*Taq 0.2μl
+
*Taq 0.2μl
 
 
 
 
  
 
==AAT 230==  
 
==AAT 230==  
Line 111: Line 108:
  
 
 
*H20 3.85μl
+
*H20 6.85μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 2.4μl
 
*MgCl2 (25mM) 2.4μl
Line 117: Line 114:
 
*230R 0.6μl
 
*230R 0.6μl
 
*230F 0.25μl
 
*230F 0.25μl
*Taq 0.2μl
+
*Taq 0.2μl
 
 
  
 
==AAT 233==  
 
==AAT 233==  
Line 142: Line 138:
 
Cycles: 25
 
Cycles: 25
  
*H20 4.45μl
+
*H20 7.45μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 1.8μl
 
*MgCl2 (25mM) 1.8μl
Line 148: Line 144:
 
*240R 0.6μl
 
*240R 0.6μl
 
*240F 0.25μl
 
*240F 0.25μl
*Taq 0.2μl
+
*Taq 0.2μl
 
 
  
 
==AAT 242==
 
==AAT 242==
Line 207: Line 202:
  
  
Annealing Temp 49
+
Annealing Temp: 49
 +
 
 +
Cycles: 30
  
*H20 3.85μl
+
*H20 6.85μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 2.4μl
 
*MgCl2 (25mM) 2.4μl
Line 238: Line 235:
  
 
Annealing Temp: 53
 
Annealing Temp: 53
 +
 
Cycles: 24
 
Cycles: 24
  
Line 243: Line 241:
  
 
 
*H20 2.95μl
+
*H20 5.95μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 1.8μl
 
*MgCl2 (25mM) 1.8μl
Line 250: Line 248:
 
*278R 0.6μl
 
*278R 0.6μl
 
*278F 0.25μl
 
*278F 0.25μl
*Taq 0.2μl
+
*Taq 0.2μl
 
 
  
 
==AAT 283==
 
==AAT 283==
Line 295: Line 292:
  
 
 
*H20 5.05μl
+
*H20 8.05μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 1.2μl
 
*MgCl2 (25mM) 1.2μl
Line 301: Line 298:
 
*300R 0.6μl
 
*300R 0.6μl
 
*300F 0.25μl
 
*300F 0.25μl
*Taq 0.2μl
+
*Taq 0.2μl
 
 
  
 
==AAT 308==
 
==AAT 308==
Line 377: Line 373:
 
Cycles: 25
 
Cycles: 25
 
 
*H20 5.05μl
+
*H20 8.05μl
 
*10X Buffer 1.5μl
 
*10X Buffer 1.5μl
 
*MgCl2 (25mM) 1.2μl
 
*MgCl2 (25mM) 1.2μl

Latest revision as of 21:37, 12 December 2006

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

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