Based on the analysis of miRNAs omics in longan ECs, we selected the differentially expressed miR396a3p_2 and miR396b5p for further analysis and verification, which play a key role in the regulatory pathway about effect of blue light on longan. We analyzed the phylogenetic tree of precursor and mature sequences, the secondary structure and cisacting element of precursor, the prediction of target genes, as well as expression patterns responding to blue light of miR396 in this study. These results showed that: (1) the miR396 mature sequences from 5p arm were highly conserved and from 3p arm were quite specific. The stem sequence was more conserved than loop sequence, and the miRNA from 5p arm sequence was more conserved than that from the 3p arm. (2) They both contain many lightresponsive elements, hormoneresponsive elements, and biotic and abiotic stress responsive elements, which might play a regulatory role in response to blue light through these ciselements. Their targets responding to blue light both contain growthregulating factor 2 (GRF2), LRR receptorlike serine/threonineprotein kinase (FLS2), ethyleneinsensitive protein 3 (EIN3) and DNAdirected RNA polymerase subunit alpha (ropA) etc. Realtime quantitative PCR showed that there were differences in the expression patterns of miR396a3p_2 and miR396b5p under different light qualities. The expression trends of miR396b5p and its targets presented negative correlation under the blue light conditions with different light intensities, while miR396a3p_2 was not negatively correlated with rpoA. This might be the reason that other miR396 family members or miRNAs were also involved in this process to achieve regulation of target gene transcription levels.