The warning comes into force at 9pm on Wednesday and will lift at 9am on Thursday.
It covers much of Dumfries and Galloway reaching Jedburgh in the east and Moffat in the north west.
A yellow warning had already been issued covering a wider area – down to Wigtown in the south west and Sanquhar in the north – but the warnings have been updated as the likelihood of severe weather in some areas increased.
The wider yellow warning is still currently in place and is due to lift at 3pm on Thursday.
The Met Office has predicted that heavy and persistent rain is likely to bring flooding and transport disruption, with spray and surface water on road leading to difficult driving conditions.
The forecaster has added that “fast flowing or deep floodwater is likely” which can present a danger to life.
Transport Scotland’s Multi-Agency Response Team, which is already operational for COP26, will monitor conditions throughout the amber warning period.
Minister for Transport Graeme Dey said: “The Met Office is warning us to expect a period of difficult weather in parts of southern Scotland, with heavy rain likely to impact travel in the amber warning area.
“The conditions could potentially bring disruption to the trunk road and rail networks, so it’s important people plan their journeys before they set off.
“Motorists should make sure their routes are available, follow travel advice from Police Scotland and drive to the conditions. The Traffic Scotland twitter page is regularly updated and the mobile website - my.trafficscotland.org - lets people access the latest information.”
Chief Superintendent Louise Blakelock has agreed and urged motorists to plan ahead and avoid travelling if at all possible.
She said: “The Met Office, SEPA and Traffic Scotland websites have valuable information about weather disruptions and we would advise people to check these sites before setting off on your journey.
“If you do need to drive, make sure you and your vehicle are suitably prepared by having sufficient fuel, supplies such as warm clothing, food and water in the event you are delayed, charge your phone and plan your route.”
Network Rail has also issued a request asking passengers to check before they travel between Wednesday and Friday this week as services on some routes will be reduced and speed limits will be in place.
Those with plans to travel between Glasgow, or Edinburgh, and Carlisle are being asked to consider if their journey is essential and only travel if unavoidable.
Speed limits will also affect services between Edinburgh and Glasgow, Edinburgh and Newcastle and the West Highland lines.
Liam Sumpter, Network Rail Scotland route director, said: “Extreme rainfall can pose a serious risk to the railway causing landslips or damaging our infrastructure and bridges. Slowing trains down will mean drivers will be better able to halt their train when a problem is spotted.
“The safety of our passengers and colleagues is our main priority during periods of poor weather and slowing services down, and running fewer trains, will help us manage these conditions for everyone.”