| It is common to receive a heterogenous population in an oligo without additional purification because of the efficiency of the coupling reaction used to product the oligos. Oligos are synthesized base by base through a series of chemical reactions. The coupling reaction itself is only about 99% efficient, so with each base addition, approximately 1% of the oligos will not pick up the proper base. We try to prevent these oligos from participating in later base additions by running through a capping reagent to block the 5' OH from coupling with a base later in the sequence. Most oligos will pick up this capping reagent and will become truncation mutants. Unfortunately, because the capping reaction is not 100% efficient, some oligos will remain uncapped and able to react with later bases. This leads to the formation of deletion mutants. This can happen anywhere within the sequence and leads to a heterogenous population. In a 27mer oligo, approximately 70% of the oligos will be full length and 30% will contain either a deletion mutation or will be truncated on the 5' end. Because of this, we strongly recommend PAGE purification for any oligo used for cloning regardless of length. This will filter out a great deal of the truncation and deletion mutants. |
|
 |
|
|
|
No Related Articles Available.
|
|
|
|
|
No Attachments Available.
|
|
|
|
|
No Related Links Available.
|
|
|
|
|
|
|
|
|
|
|