Q: For the initial submission to Nature, is it required to segregate the content into main content and supplementary information?

The guidelines for the initial submission to Nature (found here) do not explicitly say so, neither do those for the supplementary information (found here). So, you should be okay with submitting as one document, unless this is a mandatory field in the submission form. In case it’s not, you may presently still want to identify for yourself what the supplementary information is/could be, in case your manuscript is accepted. Supplementary information, as you may know, is information that is integral to the paper but is too huge or voluminous (such as a large figure/table or dataset) to be accommodated into the main paper for reasons of space. As you may understand, this may change during the course of the review and revisions, so this is probably not a critical consideration for now.

Hope that helps. For more expert help with your submission, you may also consider our submission service: Editage Journal Submission

All the best for your submission!

Related information:

• Can a supplemental material be attached to a non-open access paper?
• Guiding right: Our top 10 questions and answers on submission guidelines in 2020