You have to go back a long way! One by Phil Lawley (BBA, 21 (1956) 481-488) I have a PDF of. A summary/memoire by Rene Thomas ( Gene, 135 (1993) 77-79) contains other relevant refs. Note that after elution with pure water, partial renaturation will occrr when sodium ions are added. But my advice is to elute with the kit elution buffer, or TE, which is what that might well be.
You have to go back a long way! One by Phil Lawley (BBA, 21 (1956) 481-488) I have a PDF of. A summary/memoire by Rene Thomas ( Gene, 135 (1993) 77-79) contains other relevant refs. Note that after elution with pure water, partial renaturation will occrr when sodium ions are added. But my advice is to elute with the kit elution buffer, or TE, which is what that might well be.
If you are planning to carry out restriction digest of eluted DNA, it is not a good idea to use water: In pure water, DNA is singlestranded. This was published ages ago, but people seem to have forgotten.
If you are planning to carry out restriction digest of eluted DNA, it is not a good idea to use water: In pure water, DNA is singlestranded. This was published ages ago, but people seem to have forgotten.
During elution the DNA pellets extracted are dissolved in nuclease free water to increase recovery of the DNA TE buffer can also be used as an alternative. But the buffer used in elution should be the one used to blank the nanodrop
During elution the DNA pellets extracted are dissolved in nuclease free water to increase recovery of the DNA TE buffer can also be used as an alternative. But the buffer used in elution should be the one used to blank the nanodrop
@Geoff margison but I just googled and found that DNA will dissociate into single strand only when in poor solvent. Water is a good solvent for DNA since negative phosphate backbone hydrogen bonded with polar water molecules? Sorry if I am wrong.
@Geoff margison but I just googled and found that DNA will dissociate into single strand only when in poor solvent. Water is a good solvent for DNA since negative phosphate backbone hydrogen bonded with polar water molecules? Sorry if I am wrong.
The eluted DNA will be more stable in a buffered solution so if you intend to store the DNA for an extended period of time (e.g. weeks) you better elute using a buffer. Eluting with water or a buffer of low ionic strength may give you higher yield and also less interference and more freedom with your subsequent reactions. I personally prefer nuclease-free water.
The eluted DNA will be more stable in a buffered solution so if you intend to store the DNA for an extended period of time (e.g. weeks) you better elute using a buffer. Eluting with water or a buffer of low ionic strength may give you higher yield and also less interference and more freedom with your subsequent reactions. I personally prefer nuclease-free water.
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says - Do so at 37ºC instead of at room temperature - Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says - Do so at 37ºC instead of at room temperature - Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says - Do so at 37ºC instead of at room temperature - Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says - Do so at 37ºC instead of at room temperature - Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
Rachel Andrea Chea good luck with your cloning! Go team NTU!
Rachel Andrea Chea good luck with your cloning! Go team NTU!
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The eluted DNA will be more stable in a buffered solution , I only used Water for eluting when send for sequencing
The eluted DNA will be more stable in a buffered solution , I only used Water for eluting when send for sequencing
More
VOTE
You have to go back a long way! One by Phil Lawley (BBA, 21 (1956) 481-488) I have a PDF of. A summary/memoire by Rene Thomas ( Gene, 135 (1993) 77-79) contains other relevant refs. Note that after elution with pure water, partial renaturation will occrr when sodium ions are added. But my advice is to elute with the kit elution buffer, or TE, which is what that might well be.
You have to go back a long way! One by Phil Lawley (BBA, 21 (1956) 481-488) I have a PDF of. A summary/memoire by Rene Thomas ( Gene, 135 (1993) 77-79) contains other relevant refs. Note that after elution with pure water, partial renaturation will occrr when sodium ions are added. But my advice is to elute with the kit elution buffer, or TE, which is what that might well be.
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VOTE
If you are planning to carry out restriction digest of eluted DNA, it is not a good idea to use water: In pure water, DNA is singlestranded. This was published ages ago, but people seem to have forgotten.
If you are planning to carry out restriction digest of eluted DNA, it is not a good idea to use water: In pure water, DNA is singlestranded. This was published ages ago, but people seem to have forgotten.
More
VOTE
During elution the DNA pellets extracted are dissolved in nuclease free water to increase recovery of the DNA TE buffer can also be used as an alternative. But the buffer used in elution should be the one used to blank the nanodrop
During elution the DNA pellets extracted are dissolved in nuclease free water to increase recovery of the DNA TE buffer can also be used as an alternative. But the buffer used in elution should be the one used to blank the nanodrop
More
VOTE
@Geoff margison but I just googled and found that DNA will dissociate into single strand only when in poor solvent. Water is a good solvent for DNA since negative phosphate backbone hydrogen bonded with polar water molecules? Sorry if I am wrong.
@Geoff margison but I just googled and found that DNA will dissociate into single strand only when in poor solvent. Water is a good solvent for DNA since negative phosphate backbone hydrogen bonded with polar water molecules? Sorry if I am wrong.
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VOTE
Geoff Margison, would you mind putting up a paper reference. There aren't many publications regarding DNA interactions in pure water
Geoff Margison, would you mind putting up a paper reference. There aren't many publications regarding DNA interactions in pure water
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VOTE
The eluted DNA will be more stable in a buffered solution so if you intend to store the DNA for an extended period of time (e.g. weeks) you better elute using a buffer. Eluting with water or a buffer of low ionic strength may give you higher yield and also less interference and more freedom with your subsequent reactions. I personally prefer nuclease-free water.
The eluted DNA will be more stable in a buffered solution so if you intend to store the DNA for an extended period of time (e.g. weeks) you better elute using a buffer. Eluting with water or a buffer of low ionic strength may give you higher yield and also less interference and more freedom with your subsequent reactions. I personally prefer nuclease-free water.
More
VOTE
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says
- Do so at 37ºC instead of at room temperature
- Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
Cheers.
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says
- Do so at 37ºC instead of at room temperature
- Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
Cheers.
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Maziar S. Ardejani ah okay. You have a point. Thank you for your input!
Maziar S. Ardejani ah okay. You have a point. Thank you for your input!
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VOTE
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says
- Do so at 37ºC instead of at room temperature
- Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
Cheers.
The reason you were told to use water instead of elution buffer is that sometimes, the elution buffer has EDTA or some other substance like that. If you plan on performing ligation on such purified fragments, then you should use water instead of elution buffer because EDTA inhibits the reaction. If you are worried about the yield:
- Let it penetrate the membrane for 5 minutes instead of 1 minute as the protocol usually says
- Do so at 37ºC instead of at room temperature
- Do this twice, meaning: collect the DNA after the first centrifugation and place it back in the membrane for 5 minutes at 37ºC again.
All of this helps increase the yield. We use water instead of elution buffer all the time in our lab (because we want to ligate them to plasmids afterwards) and we are fine.
Cheers.
More
VOTE