To illustrate the mechanism of lycopene accumulation in red-fleshed Cara Cara navel orange (Citrus sinensis Os beck), the genes encoding citrus lycopene beta-cyclases (CLCYbs) were compared among citrus accessions. Protein sequence alignment showed that CLCYbs could be classified into two types, CLCYb1s and CLCYb2s, based on the polymorphisms in the 4(th) and the 50(th) amino acids. Polymerase chain reaction - Restriction fragment length polymorphisms analysis (PCR-RFLP) further indicated that all studied eighteen accessions of citrus and its relatives (including four with lycopene accumulatedflesh) could be classified into three groups. Pummelo (Citrus grandis), two grapefruits (C. paradisi), trifoliata orange (Pocirus trifoliata), two citranges (C. sinensis x P trifoliata) and kumquat (Fortunella crassifolia) are in Group-1, which are homozygous for CLCYb1s. Genomes in Group-2 and 3 are heterozygous for CLCYb1s and CLCYb2s. However, Group-2 includes two navel oranges and two sour oranges (C. aurantium), in which close amounts of Cslcyb1s and Cslcyb2s were amplified. Whereas Group-3 includes two lemons (C. limon), two mandarins (C. reticulata) and Calamondin, in which unequal and unstable levels of two variants were amplified. Southern blot analysis implied that CLCYbs had at least two copies in Cara Cara navel orange and other three tested genotypes. The data presented here indicated the irrelevancy of the two variants in their genomes to the accumulation of lycopene in the citrus fruits. Biotechnol. & Biotechnol. Eq. 2011, 25(3), 2452-2457