Difference between revisions of "ExoSAP Reaction"
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This prepares a PCR product from a [[PCR]] reaction for [[Sanger Sequencing]]. | This prepares a PCR product from a [[PCR]] reaction for [[Sanger Sequencing]]. | ||
| − | The ExoSAP reaction "removes" the | + | The ExoSAP reaction "removes" the [[dNTPs]] and [[PCR primers]] from the [[PCR product]]. It contains [[Exonuclease I]] and [[Shrimp Alkaline Phosphatase]] that chop up single stranded primers and inactivate dNTP's so they do not interfere with the sequencing reaction. The mixture is: |
:10 μl PCR product | :10 μl PCR product | ||
| − | :3 μl ExoSAP (which is 1 μl of Exonuclease I and 2 μl of Shrimp Alkaline Phosphatase) | + | :3 μl ExoSAP (which is 1 μl of [[Exonuclease I]] and 2 μl of [[Shrimp Alkaline Phosphatase]]) |
The mixture is heated to 37 C for 15 min then 80 C for 15 min. This is under the program "EXSAP" on the thermocycler. | The mixture is heated to 37 C for 15 min then 80 C for 15 min. This is under the program "EXSAP" on the thermocycler. | ||
Latest revision as of 10:13, 21 July 2018
This prepares a PCR product from a PCR reaction for Sanger Sequencing.
The ExoSAP reaction "removes" the dNTPs and PCR primers from the PCR product. It contains Exonuclease I and Shrimp Alkaline Phosphatase that chop up single stranded primers and inactivate dNTP's so they do not interfere with the sequencing reaction. The mixture is:
- 10 μl PCR product
- 3 μl ExoSAP (which is 1 μl of Exonuclease I and 2 μl of Shrimp Alkaline Phosphatase)
The mixture is heated to 37 C for 15 min then 80 C for 15 min. This is under the program "EXSAP" on the thermocycler.
The result is an "ExoSAP'd PCR product" for Sanger Sequencing
What Links Here
- Methods (← links)
- Sanger Sequencing (← links)
