Abstract: Through the method of RT-PCR, cDNA sequence of △5-fatty acid elongase gene of Phaeodactylum tricornutum Bohlin was cloned, and the double enzyme digestion technique was used to construct the recombinant expression vector, and then the related bioinformatics analysis of the gene was carried out. The results showed that the full-length of cDNA of △5-fatty acid elongase gene in Phaeodactylum tricornutum Bohlin was 834 bp, 278 amino acids were encoded, the predicted isoelectric point was 9.13, and the theoretical molecular weight was 32 348.91. There was an intron existed in △5-fatty acid elongase gene, and the enzyme belonged to ELO series elongase, which could be located in the endoplasmic reticulum and contained multiple transmembrance domains and conserved domains. The genetic relationship analysis showed that △5-fatty acid elongase of Phaeodactylum tricornutum Bohlin had the closest relationship with Thalassiosira pseudonana Hasle et Heimdal. The over-expression recombinant plasmid pPha-T1-5e of △5-fatty acid elongase gene in Phaeodactylum tricornutum Bohlin was also successfully constructed in the experiment, which laid a foundation for the expression and functional verification of related lipid metabolism genes in Phaeodactylum tricornutum Bohlin.