A length polymorphism in the trnL/trnF intergenic spacer was used as a marker to determine the mode of chloroplast inheritance in Streptocarpus (Gesneriaceae). Exclusively maternal inheritance was recorded for all the F1 progeny of reciprocal intraspecific crosses between S. primulifolius and a population referred to as S. aff. primulifolius from the Igoda River mouth, Eastern Cape, South Africa, and for interspecific crosses between S. rexii and S. dunnii. A combination of molecular and morphological data was used to clarify the origin of S. aff. primulifolius, which possesses S. rexii-type cpDNA and rDNA, while the morphological data suggest an intermediate position between S. rexii and S. primulifolius. The distribution of S. rexii and S. primulifolius, combined with molecular and morphological data, supports the hypothesis that the S. aff. primulifolius population is a hybrid between S. rexii and S. primulifolius, with S. rexii as the maternal parent, and that substantial molecular but limited morphological introgression into S. primulifolius has taken place.