Predicting H3K27M mutation status in midline gliomas non-invasively is of considerable interest, particularly using deep learning with 11C-methionine (MET) and 18F-fluoroethyltyrosine (FET) positron emission tomography (PET). To optimise prediction efficiency, we derived an assistance training (AT) scheme to allow mutual benefits between MET and FET learning to boost the predictability but still only require either PET as inputs for predictions. Our method significantly surpassed conventional convolutional neural network (CNN), radiomics-based, and MR-based methods, achieved an area under the curve (AUC) of 0.9343 for MET, and an AUC of 0.8619 for FET during internal cross-validation (
n
= 90). The performance remained high in hold-out testing (
n
= 19) and consecutive testing cohorts (
n
= 21), with AUCs of 0.9205 and 0.7404. The clinical feasibility of the proposed method was confirmed by the agreements to multi-departmental decisions and outcomes in pathology-uncertain cases. The findings positions our method as a promising tool for aiding treatment decisions in midline glioma.