Why is the cloning step unnecessary prior to sequencing with massively parallel sequencing (MPS; Illumina or similar technologies), when it is necessary for hierarchical and shotgun sequencing?

I know that the cloning step is helpful for introducing universal primer binding sites for sequencing. So is it because we are using probes with adaptors on them for MPS that we don't need to clone?

  • $\begingroup$ Can you clarify what MPS is or link to some information that describes in more detail what you mean? I can imagine that "MPS" is "massively parallel sequencing" but that acronym is not standard terminology, as far as I know. $\endgroup$ Commented Apr 19, 2022 at 17:01
  • 1
    $\begingroup$ @MaximilianPress Oh sorry, yes I mean massively parallel sequencing. $\endgroup$
    – katara
    Commented Apr 19, 2022 at 17:14

1 Answer 1


The construction of sequencing libraries for Illumina, PacBio, ONT, or similar instruments is a large field of methodology that encompasses a variety of different techniques. For background on Illumina libraries specifically you can see here.

In some cases, notably Illumina, indeed the library construction involves sequencing adapters either being ligated or added by PCR. There are many strategies here and I don't claim exhaustive knowledge of all of them, I would suggest doing more research here if you are interested.

For other cases, sequencing occurs by alternative mechanisms. I suggest consulting overviews of PacBio and nanopore (ONT) sequencing to get ideas of how different methods work for the same basic task of shotgun sequencing of DNA without a cloning step.


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