Potato yellow vein virus (PYVV) with tripartite RNA (ss+) genome is classified as Crinivirus within the family Closteoviridae. PYVV is the causal agent of potato yellow vein disease (PYVD) with yield effect. Single strand conformational polymorphism (SSCP) was reported to estimate variability in eukaryotes and different viruses’ species. In this study, the molecular variability of PYVV was analyzed by SSCP patterns regarding three genes: major capsid protein (CP), minor capsid protein (CPm), and heat shock protein (Hsp70) obtained from isolates of 60 potato plants leaves expressing PYVD. Leaves were collected in Nariño, Colombia from 30 Solanum tuberosum, Phureja Group (PhG) and 30 Andigena Group (AG). Genes were amplified by RT-PCR and the purified direct PCR products were used for SSCP. Three SSCP patterns were detected for CP gen, 12 for CPm and 12 for Hsp70 genes. The AG host, most frequent pattern was C (66.6%) for the Hsp70 gene and IV (33.3%) for CPm gene. Complex SSCP patters with inter and intra variation were detected. CP gene pattern 1 was present in 93.3% of the isolates of both host groups, indicating low number of variants compared to CPm and Hsp70 genes. This is the first attempt to estimate intra e inter PYVV variability by the use of a simple molecular method considering three genes in a large group of potato samples affected by PYVD. SSCP technique was useful to discriminate the migration patterns generated for each gene