Among the tools of modern biotechnologies tissue culture is being increasingly applied to complement other in-situ or ex-situ conservation methods. Within the Institute of Biology from Bucharest, several in vitro techniques have been developed, with recent establishment of endangered plant tissue culture collections. Hormonal regulation of the regeneration process was histologically monitored in order to develop for the first time, a protocol for efficient multiplication and establishment of an in vitro collection of Alyssum borzaeanum germplasm. The most efficient hormonal combination for shoot regeneration was 4.43 μM BAP and 0.57 μM IAA. The described correlations between morphogenetic pattern and growth regulators type and content enable the control upon the development processes being useful for further investigation and use of Alyssum species.