The botanical garden was founded in 1810, during the Illyrian Provinces, as the Garden of Native Flora as part of the then high school (Écoles Centrales). It was designed by Franc Hladnik (1773–1844), who was its first director and was also a lecturer on natural science and botany at the above school. Just how important the garden was to the French authorities is shown by the fact that one of the first trees, a linden tree, was planted by Marshall Marmont, the former head governor of the Illyrian Provinces.
The garden initially covered an area of 33 ares, it received an annual subsidy of 1,000 francs and had a gardener employed who earned an annual salary of 500 francs. For the year 1812, Hladnik lists 768 native plant species, which means that the garden developed rapidly under the French rule. It is thanks to Hladnik and his links with Austrian botanists that after the reinstatement of Austrian rule the garden retained its unchanged form. After 1822, it was twice enlarged. With the founding of the University of Ljubljana, the garden came under its patronage in 1920 and even today continues to be part of the Department of Biology of the Biotechnical Faculty. In 2008, it gained the status of a monument of national importance.
The Ljubljana Botanical Garden has carried on its activities in the same location for almost 200 years and it is therefore one of the most important old European botanical gardens.
The institution is a member of the international network Botanic Gardens Conservation International and cooperates with more than 270 botanical gardens all across the world. Of over 4,500 plant species and subspecies that grow on 2 hectares (4.9 acres), roughly a third is endemic to Slovenia, whereas the rest originate from other European places and other continents.