Restriction fragment length polymorphism (RFLP) analysis of PCR products amplified from 18S ribosomal RNA gene of Trypanosoma congolense

Abstract

Oligonucleotide primers were designed from the conserved nucleotide sequences of 18S ribosomal RNA (18S rRNA) gene of protozoans: Trypanosoma brucei, Leishmania donooani, Triponema aequale and Lagenidium gigantum. The primers were used in polymerase chain reaction (PCR) to generate PCR products of approximately 1 K.b using genomic DNA from T.brucei and the four genotypic groups of T.congolense as template. The five PCR products so produced were digested with several restriction enzymes and hybridized to a DNA probe made from Tsbrucei PCR product of the same 18S rRNA gene region. Most restriction enzyme digests revealed polymorphism with respect to the location of their recognition sites on the five PCR products. The restriction fragment length polymorphism (RFLP) pattern observed indicate that the 18S rRNA gene sequences of trypanosomes: T.brucei and the four genotypes of T.congolense group are heterogeneous. The results further demonstrate that the region that was amplified can be used in specific identification of trypanosome species and subspecies.