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Team:Newcastle/Gel extraction
From 2010.igem.org
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=QIAquick Gel Extraction Microcentrifuge Protocol= | =QIAquick Gel Extraction Microcentrifuge Protocol= | ||
| - | # Excise the DNA fragment from the agarose gel with a clean, sharp scalpel | + | # Excise the DNA fragment from the agarose gel with a clean, sharp scalpel. |
| - | # Weight the gel slice and add 3 volumes of buffer QG to 1 volume of gel (100 mg ~ 100 µl) | + | # Weight the gel slice and add 3 volumes of buffer QG to 1 volume of gel (100 mg ~ 100 µl). |
| - | # Incubate at 50°C and invert the tube gently at regular | + | # Incubate at 50°C and invert the tube gently at regular interval until the gel has completely dissolved. |
| - | # After the gel has | + | # After the gel has dissolved completely, check that the color of the mixture is yellow. |
| - | # Add 1 gel volume of isopropanol to the sample and mix | + | # Add 1 gel volume of isopropanol to the sample and mix. |
# Place a QIAquick spin column in a 2 ml collection tube | # Place a QIAquick spin column in a 2 ml collection tube | ||
| - | # To bind DNA, | + | # To bind DNA, apply the sample to the QIAquick column and centrifuge for 1 min. |
| - | # Discard the flow through and place the QIAquick column back into the same tube ( | + | # Discard the flow through and place the QIAquick column back into the same tube (max volume: 750 µl). |
| - | # Add 0.5 ml of Buffer QG to QIAquick column and centrifuge for 1 min | + | # Add 0.5 ml of Buffer QG to QIAquick column and centrifuge for 1 min. Discard the flow through and place the QIAquick column back into the same tube. |
| - | # To wash, add 0.75 ml of Buffer PE to QIAquick column and centrifuge for 1 min | + | # To wash, add 0.75 ml of Buffer PE to QIAquick column and centrifuge for 1 min. Place the QIAquick column back into the same tube. |
| - | # Centrifuge the column for a further 1 min | + | # Centrifuge the column for a further 1 min. |
| - | # Transfer the column into a clean 1.5 ml micriocentrifuge tube | + | # Transfer the column into a clean 1.5 ml micriocentrifuge tube. |
| - | # To elute DNA, add 30 µl of Buffer EB to the center of the QIAquick membrane and allow to stand for 1 min | + | # To elute DNA, add 30 µl of Buffer EB to the center of the QIAquick membrane and allow to stand for 1 min. |
| - | # Centrifuge the column for 1 min and transfer the eluate to a clean 1.5 ml micriocentrifuge tube | + | # Centrifuge the column for 1 min and transfer the eluate to a clean 1.5 ml micriocentrifuge tube. |
{{Team:Newcastle/footer}} | {{Team:Newcastle/footer}} | ||
Revision as of 09:06, 10 August 2010
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QIAquick Gel Extraction Microcentrifuge Protocol
- Excise the DNA fragment from the agarose gel with a clean, sharp scalpel.
- Weight the gel slice and add 3 volumes of buffer QG to 1 volume of gel (100 mg ~ 100 µl).
- Incubate at 50°C and invert the tube gently at regular interval until the gel has completely dissolved.
- After the gel has dissolved completely, check that the color of the mixture is yellow.
- Add 1 gel volume of isopropanol to the sample and mix.
- Place a QIAquick spin column in a 2 ml collection tube
- To bind DNA, apply the sample to the QIAquick column and centrifuge for 1 min.
- Discard the flow through and place the QIAquick column back into the same tube (max volume: 750 µl).
- Add 0.5 ml of Buffer QG to QIAquick column and centrifuge for 1 min. Discard the flow through and place the QIAquick column back into the same tube.
- To wash, add 0.75 ml of Buffer PE to QIAquick column and centrifuge for 1 min. Place the QIAquick column back into the same tube.
- Centrifuge the column for a further 1 min.
- Transfer the column into a clean 1.5 ml micriocentrifuge tube.
- To elute DNA, add 30 µl of Buffer EB to the center of the QIAquick membrane and allow to stand for 1 min.
- Centrifuge the column for 1 min and transfer the eluate to a clean 1.5 ml micriocentrifuge tube.
   
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