Euphytica 116: 243–250, 2000. © 2000 Kluwer Academic Publishers. Printed in the Netherlands. 243 PCR-RFLP analysis of chloroplast gene regions in Cajanus (Leguminosae) and allied genera Mukkamala Lakshmi, Padmanabhan Senthilkumar, Madasamy Parani, Mundaya Naray- anan Jithesh & Ajay Kumar Parida ∗ M. S. Swaminathan Research Foundation, III Cross Street, Institutional Area, Taramani, Chennai 600 113, India ( ∗ author for correspondence; phone: +91 44 235 1229; fax: +91 442 35 1319; e-mail: MDSAA51@giasmd01.vsnl.net.in) Received 6 October 1999; accepted 29 March 2000 Key words: Cajanus, Chloroplast DNA, leguminosae, PCR-RFLP, Pigeon pea, Phylogeny, Rhynchosia Summary Twenty-eight species belonging to five genera of the sub tribe Cajaninae; viz., Cajanus (15 species), Rhynchosia (10 species), Dunbaria, Flemingia and Paracalyx were analysed for variations in four chloroplast gene regions, rbcL, trnS-psbC, 16S and trnL-UAA. The four chloroplast gene regions were amplified with specific primers and subsequently digested with 15 restriction enzymes. Rhynchosia did not show any inter-specific differences in the four gene regions for any of the enzymes used. In Cajanus, the inter-specific PCR-RFLP profile of the four gene regions for all the enzymes were similar except for the Pst I digests of trnS-psbC gene region. At inter-generic level, rbcL gene region did not show polymorphism with any of the enzymes used; while in trnS-psbC gene region polymorphism was observed only with Pst I and Hae III digestions. Inter-generic PCR-RFLP of 16S did not reveal any variation. The trnL-UAA gene region had restriction site only for EcoR I in which Cajanus and Rhynchosia showed similar profiles while Dunbaria showed a different profile. The trnL-UAA gene region in Paracalyx and Flemingia did not have restriction site for this enzyme. Despite the analyses of four gene regions using 15 restriction enzymes, differentiation at genus level could not be obtained. These observations indicated limited divergence of the chloroplast genome among the four genera of the sub-tribe Cajaninae suggesting close relationships of the taxa. Introduction Pigeon pea, Cajanus cajan (L.) Millsp. is one of the major grain legume crops of the tropics and the sub-tropics. Though it is perennial, it is generally cultivated as a annual crop. The landraces and the traditional cultivars of this species which are long dur- ation crops, are also grown as inter-crops with shorter duration crops. The center of origin of this species has been very much debated as to being India or Africa. Based on the chromosome number and floral mor- phology, it was concluded by van der Maesen (1980) that pigeon pea is of Indian origin with Africa being the secondary center of diversity. The cultivated pi- geon pea has many wild relatives which belong to six genera – Cajanus, Dunbaria, Flemingia, Paracalyx, Rhynchosia and Eriosema; all of which are grouped under the sub-tribe Cajaninae most of them recorded as of Indian origin (van der Maesen, 1980). Many of the wild species belonging to Cajanus and Rhynchosia have been utilized by various workers for inter-specific hybridisation with a view to transfer de- sirable genes to the cultivated species. However, stud- ies on the genetic relationships within and between the five genera of the tribe Cajaninae have been very lim- ited. Use of molecular tools to study the phylogeny, evolutionary trends and species relationship in Ca- janus has received little attention. There are only a few reports (Ratnaparkhe et al., 1995; Nadimpalli et al., 1992) until now that attempt to address the phylogeny of the species groups, though the need to understand these aspects has been emphasised a number of times (Kumar et al., 1958; Pundir & Singh, 1985a, 1985b).