Yes, you can.
In short, yes, you can use a hardware wallet that you're already using for singlesig bitcoin custody when creating an Unchained vault.
A singlesig wallet will have a specific set of addresses derived from its unique private key.
When creating a 2-of-3 multisig wallet, your device’s public key will combine with the public key of two other devices to construct a new multisig wallet. The singlesig wallet is not even aware that it is also part of a multisig wallet.
Rest assured that the existing singlesig wallet funds will not be affected if the same extended public key is used to construct a multisig wallet.