The Gloucester centre arrived in Sydney from Argentina on Friday and will be one of three new boys thrust into the firing line for the Lions in the Australian capital on Tuesday night as the tourists try to maintain their winning momentum four days out from the first Test.
Fellow English internationals Christian Wade and Brad Barritt will join Twelvetrees in donning the famous red jersey for the first time after just one training session, and the trio will be joined by the 36-year-old Shane Williams in one of the most make-shift back lines the Lions have ever assembled.
But with Jamie Roberts and Manu Tuilagi both struggling with injuries, Tuesday night's exercise is more than just about filling-in for Twelvetrees. If he makes as big an impact as he has done before on his first outings then a Test place could be up for grabs.
Having cut his teeth at Pullborough RFC and then Haywards Heath, Leicester Lions and Nottingham, Twelvetrees made his first big impact on loan at Bedford Blues. Four tries on his debut against Manchester in 2008 marked him out from the rest and he ended that season as joint top try scorer on 18 in the Championship.
That form earned him a recall to Leicester Tigers, where he turned in a match-winning performance on his Heineken Cup debut for the club after a mere 10 minutes notice he would be playing. Dan Hipkiss pulled out in the warm-up and Twelvetrees went on to score a man of the match winning 14 points against the Ospreys.
Fast-forward to 2013, and his England and RBS 6 Nations debut, and there was a try at Twickenham in the Calcutta Cup win over Scotland to add to his dream debuts. So what can we all expect from him on his Lions debut?
"I want to take the chance that has been offered to me with both hands, play some good rugby and win with the Lions. Everyone here sets very high standards and I want to contribute to the side," said Twelvetrees.
"From my experience of being here for two days you are buzzing on adrenalin because you are here playing for the Lions. It's a very exciting thing and we are all honoured to be here and can't wait to get the shirt on.
"I got here on Friday and couldn't wait to start training the next day. It is a pretty awesome feeling to be called-up by the Lions because a few days ago I was concentrating on trying to win a series in Argentina with England - it's surreal!
"You've got to believe in your own ability - that's what has got me where I am this year. From joining Gloucester, to starting many games in the Premiership, to making a debut for England and now being here with the Lions.
"You make of it what you want - the sky is the limit. I've tried hard all year and got feedback off the coaches and now I have been given the opportunity to play for the Lions."
Twelvetrees will have the man who partnered him in the midfield on his England debut, Barritt, alongside him in Canberra on Tuesday night and his former Leicester Tigers team mate Ben Youngs not far away at scrum half.
The Brumbies have been the stand-out team in the Australian Super Rugby Conference this season and are already in the play-offs. They will fancy their chances of upsetting a Lions side featuring so many new players, but that doesn't faze Twelvetrees.
"All the players here set very high standards within themselves. Within the squad we push ourselves and that showed in the game on Saturday night against the Waratahs," he added.
"People want to play good rugby and winning rugby and I want to be there contributing to the side, otherwise there is no point in being here. Rugby is all to do with confidence and your mentality.
"If you think you can do it, you are going to do it most of the time. You have to back your ability. That's what's got me here - playing regular rugby and backing myself.
"It's a tough environment to be in and you have to have the right mentality to take it all the way."