Stena currently operates from Stranraer but has been discussing moving closer to the mouth of Loch Ryan since 2003.
"The relocation of the port to the new Loch Ryan Port will aid tourism with faster crossings between Scotland and Northern Ireland and help the regeneration of Stranraer as a major marine leisure area," said First Minister of Scotland Alex Salmond.
The investment will include the addition of two large modern ferries when the port becomes fully operational on its Irish Sea service to Belfast Port.
"This new development will safeguard and create 1,400 jobs and trigger significant investment in Dumfries and Galloway," added Salmond.
The creation of the new port will aid the regeneration of the waterfront at Stranraer, providing an opportunity to develop 26 acres of waterfront and strengthen the town's reputation as a marine leisure destination.
"This is an historic day for the people of Scotland and indeed Northern Ireland. The investment we have announced will provide Scotland with a first class port facility which underlines and enhances the port's prominence as the third largest gateway in the UK," said Dan Sten Olsson, chairman of Stena Line.
Work at the new port is expected to take around 20 months to complete.
Stena Line operates in Scandinavia, the North Sea and the Irish Sea with a network of 18 ferry routes in Europe.