First of all, determining the purification and accurate quantity of short PCR products is very difficult and the general PCR products purification kit require products to be longer than 150bp, therefore we need the PCR products used for sequencing to be no less than 150bp in length. Secondly, it is owing to the limitation of sequencing itself. For example, sequencing a 100bp PCR product with the sequence of two primers about 40 to 50bp and some unreadable bases at the beginning of the sequencing being removed, the useful sequence that can be really obtained is no more than 30-40 bases. Therefore, the short PCR product can only be cloned and then sequenced.
Articles in this section
- Will the primers degrade at room temperature during transportation? Can we use dry ice for transporting to maintain stability?
- Why there is a difference between my sequencing data and the one from the references?
- Why the short PCR product is not suitable for direct sequencing?
- Why target gene products are seen in control sample with no template?
- Why my sequence results are reverse complement to the template DNA, while I demand a 5’ to 3’ orientation?
- Why is the TM value for primers of Genscript different from the TM value I obtained from other software?
- Why is the sequence of primers in the sequencing results different from the expected sequence?
- Why is my gene sequence different from the standard sequence?
- Why I got a sequencing result that was not I want?
- Why does the sequencing primer need to be site-specific on the DNA template?