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Minnesota/27 July 2008
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|'''1. Spec the plasmid prep products:''' Place 10 samples into 10 separate plate wells. 11 wells total were used: 1st well was the control so it had 36uL of distilled water and 4uL of elution buffer (since want everything except DNA to be control group), then 2-11 wells had 1-10 plasmid prep products. 2-11 wells had 36uL of distilled water added and 4uL of plasmid prep products. Results: | |'''1. Spec the plasmid prep products:''' Place 10 samples into 10 separate plate wells. 11 wells total were used: 1st well was the control so it had 36uL of distilled water and 4uL of elution buffer (since want everything except DNA to be control group), then 2-11 wells had 1-10 plasmid prep products. 2-11 wells had 36uL of distilled water added and 4uL of plasmid prep products. Results: | ||
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| - | |'''a.''' (1) Lac I #1 = [DNA] ng/uL is 45, [DNA] ug/uL is 0.045 HIGH/GOOD | + | |'''a.''' (1) Lac I #1 = [DNA] ng/uL is 45, [DNA] ug/uL is 0.045 ''HIGH/GOOD'' |
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| - | |'''b.''' (2) Lac I = [DNA] ng/uL is 35, [DNA] ug/uL is 0.035 HIGH/GOOD | + | |'''b.''' (2) Lac I = [DNA] ng/uL is 35, [DNA] ug/uL is 0.035 ''HIGH/GOOD'' |
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| - | |'''c.''' (3) Lac I = [DNA] ng/uL is 35, [DNA] ug/uL is 0.035 HIGH/GOOD | + | |'''c.''' (3) Lac I = [DNA] ng/uL is 35, [DNA] ug/uL is 0.035 ''HIGH/GOOD'' |
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| - | |'''d.''' (4) BV:Lac #2 = [DNA] ng/uL is 10, [DNA] ug/uL is 0.001 LOW/POOR | + | |'''d.''' (4) BV:Lac #2 = [DNA] ng/uL is 10, [DNA] ug/uL is 0.001 ''LOW/POOR'' |
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| - | |'''e.''' (5) BV:Tet #1 = [DNA] ng/uL is 0, [DNA] ug/uL is 0.00 | + | |'''e.''' (5) BV:Tet #1 = [DNA] ng/uL is 0, [DNA] ug/uL is 0.00 '' NONE/BAD'' |
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| - | |'''f.''' (6) BV:Tet #2 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 | + | |'''f.''' (6) BV:Tet #2 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 '' LOW/POOR'' |
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| - | |'''g.''' (7) BV:Tet/p22 #1 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 | + | |'''g.''' (7) BV:Tet/p22 #1 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 '' LOW/POOR'' |
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| - | |'''h.''' (8) BV:Tet/p22 #1 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 LOW/POOR | + | |'''h.''' (8) BV:Tet/p22 #1 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 ''LOW/POOR'' |
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| - | |'''i.''' (9) BV:Tet/p22 #2 = [DNA] ng/uL is 335, [DNA] ug/uL is 0.335 HIGH/GOOD | + | |'''i.''' (9) BV:Tet/p22 #2 = [DNA] ng/uL is 335, [DNA] ug/uL is 0.335 ''HIGH/GOOD'' |
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| - | |'''j.''' (10) BV:Tet/p22 #2 = [DNA] ng/uL is 15, [DNA] ug/uL is 0.015 LOW/POOR | + | |'''j.''' (10) BV:Tet/p22 #2 = [DNA] ng/uL is 15, [DNA] ug/uL is 0.015 ''LOW/POOR'' |
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|'''2. Digest Rxn:''' Using #'s: 1, 2, 6, 9, and 10 from above. Also using: p22cII, LAMBDAcI, and RFP from different plasmid prep days. After following table below, allow to incubate for 2 hours. Heat inactivate enzymes @ 65C for 15 minutes in water bath. Follow the table below: | |'''2. Digest Rxn:''' Using #'s: 1, 2, 6, 9, and 10 from above. Also using: p22cII, LAMBDAcI, and RFP from different plasmid prep days. After following table below, allow to incubate for 2 hours. Heat inactivate enzymes @ 65C for 15 minutes in water bath. Follow the table below: | ||
Latest revision as of 20:52, 29 July 2008
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| 1. Spec the plasmid prep products: Place 10 samples into 10 separate plate wells. 11 wells total were used: 1st well was the control so it had 36uL of distilled water and 4uL of elution buffer (since want everything except DNA to be control group), then 2-11 wells had 1-10 plasmid prep products. 2-11 wells had 36uL of distilled water added and 4uL of plasmid prep products. Results: | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| a. (1) Lac I #1 = [DNA] ng/uL is 45, [DNA] ug/uL is 0.045 HIGH/GOOD | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| b. (2) Lac I = [DNA] ng/uL is 35, [DNA] ug/uL is 0.035 HIGH/GOOD | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| c. (3) Lac I = [DNA] ng/uL is 35, [DNA] ug/uL is 0.035 HIGH/GOOD | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| d. (4) BV:Lac #2 = [DNA] ng/uL is 10, [DNA] ug/uL is 0.001 LOW/POOR | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| e. (5) BV:Tet #1 = [DNA] ng/uL is 0, [DNA] ug/uL is 0.00 NONE/BAD | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| f. (6) BV:Tet #2 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 LOW/POOR | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| g. (7) BV:Tet/p22 #1 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 LOW/POOR | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| h. (8) BV:Tet/p22 #1 = [DNA] ng/uL is 5, [DNA] ug/uL is 0.005 LOW/POOR | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| i. (9) BV:Tet/p22 #2 = [DNA] ng/uL is 335, [DNA] ug/uL is 0.335 HIGH/GOOD | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| j. (10) BV:Tet/p22 #2 = [DNA] ng/uL is 15, [DNA] ug/uL is 0.015 LOW/POOR | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| 2. Digest Rxn: Using #'s: 1, 2, 6, 9, and 10 from above. Also using: p22cII, LAMBDAcI, and RFP from different plasmid prep days. After following table below, allow to incubate for 2 hours. Heat inactivate enzymes @ 65C for 15 minutes in water bath. Follow the table below:
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| 3. Vector Dephosphorylate: Dephosphorylate the base vectors of the digested products. After following the table below, allow to incubate for 30 minutes @37C. Place samples in water bath @ 65C to deactivate the enzyme. Place samples in freezer @ -20C O/N to continue Ligation. Follow the table below:
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