Cassava (Manihot esculenta Crantz), is an important root crop in most Neotropical countries and is one of the richest carbohydrate sources for millions of people in the tropics. This crop has a diversity of different varieties and/or cultivars with a variety of morphological, agronomic and industrial characteristics. To elucidate the genetic differentiation in cassava varieties, recent progress has been made with the application of Simple Sequence Repeats (SSR). In the present study, we characterized the genetic diversity of an in vitro Venezuelan germplasm cassava bank, located at Instituto de Estudios Avanzados, using the SSR described by CIAT (Mba et al., 2001). A total of 20 SSR markers were amplified using genomic DNA isolated from fully in vitro plants of 100 cassava accessions. In preliminary analysis of 7 SSR (SSR 12, 51, 63, 102, 103, 105, 161) a total of 33 alleles (between 2 and 7 alleles per locus) were found with a high level of polymorphism. The genetic diversity value (H) was 0.68. The highest Polymorphism Information Content (PIC) was recorder by de SSR 12 (0.82) and the lowest PIC by SSR 102 (0.38), hence the SSR 12 was found to be highly informative followed by SSR 51 (0.78). With this preliminary study, we can confirm that SSR markers can contribute to a better understanding of the genetic diversity present in this Venezuelan collection of cassava in vitro plants