I used oligos for cloning, sequenced a clone and found a mutation within the oligo sequence. Can mutations happen in synthetic oligos?
Mutations or alternations in sequence are rare in synthetic oligos but they can happen. Oligos may become depurinated in some positions during the synthesis process. The longer the oligonucleotide, the more depurinated sites may occur. These depurinations may become visible in applications like cloning where single molecules are selected and propagated. Typically the depurinated sites are replaced by any base. Usually only some oligonucleotide molecules are affected. If you have sequenced only one clone and found a ‘mutation’ in it, sequencing another independent clone will in most cases result in the correct sequence. When choosing clones, it is most important to select independent ones: To get independent clones, the pre-incubation time before plating should be based on the manufacturer instructions of your cloning system. Optimal density of the clones on the plate ensures that well separated clones can be selected. Longer oligos used for cloning should be purified to remove any trun
Related Questions
- I used an oligonucleotide in a cloning experiment an when I sequence through the area represented by the oligo the sequence is different from the sequence I ordered. Why?
- I used oligos for cloning, sequenced a clone and found a mutation within the oligo sequence. Can mutations happen in synthetic oligos?
- What type of mutation could result from a deletion of a single nucleotide from the DNA sequence?
