Rashford, who had come on against Brazil at the weekend, was uninvolved in the 2-2 draw against European opposition.
Many players on the fringes of the team got a run out, such as Ezri Konsa, Kobbie Mainoo, Jarrod Bowen, and Ivan Toney.
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On Rashford not playing due to injury, Southgate said: No, I wanted to see Anthony Gordon again.
I thought he had an excellent impact in the first game and I thought that (James) Maddison coming into that area could open things up for us a bit."
The forward will be hoping his form at United between now and the end of the season can keep him in the Euros squad for the summer.