To reveal the function of diacylglycerol acyltransferase (DGAT) on algae lipid production, DGAT from Chlorella variabilis NC64A was cloned and expressed by E.coli BL21 (DE3). The results showed that the DGAT gene was 894 bp and was coded 297aa. The apparent molecular weight of DGAT was 33 kDa and the pI was 9.48. Conserved domain analysis showed that it belonged to the Lysophospholipid acyltransferases (LPLATs) super family and the amino acids of H68, L71, F76, R94, I97 and GAA (144–146) could form special acyl acceptor binding pocket to bind acyl of ACP or CoA and served as the catalyst in the synthesis of TAG. Sequence analysis showed that DGAT shared a 32% and 24% homology with SsPDAT and AtPDAT respectively. Thin layer chromatography showed that DGAT had PDAT enzyme activity, which may promote the membrane lipid degradation and couple TAG synthesis under nitrogen starvation.