The InBIO Barcoding Initiative (IBI) Diptera 02 dataset contains records of 412 crane fly specimens belonging to the Diptera families: Limoniidae, Pediciidae and Tipulidae. This dataset is the second release by IBI on Diptera and it greatly increases the knowledge on the DNA barcodes and distribution of crane flies from Portugal. All specimens were collected in Portugal, including six specimens from the Azores and Madeira archipelagos. Sampling took place from 2003 to 2019. Specimens have been morphologically identified to species level by taxonomists and belong to 83 species in total. The species, represented in this dataset, correspond to about 55% of all the crane fly species known from Portugal and 22% of crane fly species known from the Iberian Peninsula. All DNA extractions and most specimens are deposited in the IBI collection at CIBIO, Research Center in Biodiversity and Genetic Resources.Fifty-three species were new additions to the Barcode of Life Data System (BOLD), with another 18 species' barcodes added from under-represented species in BOLD. Furthermore, the submitted sequences were found to cluster in 88 BINs, 54 of which were new to BOLD. All specimens have their DNA barcodes publicly accessible through BOLD online database and its collection data can be accessed through the Global Biodiversity Information Facility (GBIF). One species, Gonomyia tenella (Limoniidae), is recorded for the first time from Portugal, raising the number of crane flies recorded in the country to 145 species.