Team:Newcastle/Project/Labwork/OurProtocols/PCR Products

From 2009.igem.org

(Difference between revisions)
Line 8: Line 8:
This step will will be useful to determine how much of the product to use for the ligation
This step will will be useful to determine how much of the product to use for the ligation
*Digest the PCR product
*Digest the PCR product
-
Although the total will be 50ul, some of it will be consumed for the gel and to measure the concentration
+
Although the total will be 50ul, some of it will be consumed for the gel and to measure the concentration. Make a total solution of 70ul using the followings. (Check the restriction sites and replace EcoRI and SpeI as you need)
 +
- 50ul of the PCR product
 +
- 3ul of EcoRI
 +
- 3ul of SpeI
 +
- 7ul of 10X buffer
 +
- Use water to make the final volume 70ul
 +
 
 +
Mix everything well and incubate at 37C for an hour
-
Make a total solution of 70ul using the followings. (Check the restriction sites and replace EcoRI and SpeI as you need)
 
-
**50ul of the PCR product
 
-
**3ul of EcoRI
 
-
**3ul of SpeI
 
-
**7ul of 10X buffer
 
-
**Use water to make the final volume 70ul
 
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Revision as of 14:28, 1 September 2009

  • Use PCR Clean-up kit to clear the PCR product

This should result in 50ul of final product

  • Measure the concentration of the product

This step will will be useful to determine how much of the product to use for the ligation

  • Digest the PCR product

Although the total will be 50ul, some of it will be consumed for the gel and to measure the concentration. Make a total solution of 70ul using the followings. (Check the restriction sites and replace EcoRI and SpeI as you need) 

- 50ul of the PCR product
- 3ul of EcoRI
- 3ul of SpeI
- 7ul of 10X buffer
- Use water to make the final volume 70ul

Mix everything well and incubate at 37C for an hour




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