I ran a pcr using degen primers and cloned my pcr products. Because i used degen primers, there will be a few different sequences in my product (my interest gene is from a superfamily), say: sequence A, B, and C, that are 50% similar in sequence and almost the same length.
After cloning, will each colony contain only 1 of 3 types of sequence? How does this happen?
Or A,B,C can present in a single colony? IF this is the case, how should i sequence it??
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Can a single colony contains more than 2 types of insert in their plasmids?
Started by hianghao, Sep 21 2010 03:23 AM
2 replies to this topic
#2
Curtis
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Posted 21 September 2010 - 07:37 AM
it took me couple of minutes to think about your question but I couldn't come up with a good answer, it is a very technical question. However any colony grows from a single transformed bacteria. To me it sounds very unlikely to have A, B and C in the same colony.
just grow the colony in broth, extract with a reliable extraction kit and send for sequencing.
just grow the colony in broth, extract with a reliable extraction kit and send for sequencing.
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hianghao
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Posted 21 September 2010 - 08:30 AM
Curtis, on 21 September 2010 - 07:37 AM, said:
it took me couple of minutes to think about your question but I couldn't come up with a good answer, it is a very technical question. However any colony grows from a single transformed bacteria. To me it sounds very unlikely to have A, B and C in the same colony.
just grow the colony in broth, extract with a reliable extraction kit and send for sequencing.
just grow the colony in broth, extract with a reliable extraction kit and send for sequencing.
But if the colony contains 2 sequence, its quite impossible to sequence the 2 sequences from it, right? If i am not wrong, E coli is able to pick up more than 1 plasmid so why it is unlikely to have A, B and C in the same colony?
