A species specific satellite DNA family in the genome of the(2)

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O. RANDIG et al.

and Sau3A I), the presence of a strong band of≈ 270 bp was observed after electrophoresis (data not shown). The≈ 270 bp Bgl II band, which was thought to correspond to the monomeric unit of a satellite DNA family, was puri ed from the gel, 32Plabelled and used as a probe in the Southern blot analysis. The autoradiograph revealed the presence of a prominent≈ 270 bp repeat in the Alu I, Bgl II, Rsa I and Sau3A I digests, that could perhaps correspond to the monomer (Fig. 1). Strong hybridization signals of a smaller size were obtained from the Rsa I and Sau3A I digests. In the Bgl II digest, a fragment of≈ 550 bp was also detected, that could correspond to the dimer. In the Alu I digest, a ladder pattern of≈ 270 bp putative multimers was observed (Fig. 1). To further examine the arrangement of these sequences, DNA fragments produced after a time-course digestion of M. exigua DNA with Bgl II were hybridized with the probe described above. Autoradiography showed a typical ladder pattern, with an increasing amount of the monomer during digestion (Fig. 2), which is characteristic of repeated sequences arranged in tandem arrays. After complete digestion, monomers and dimers were present, which indicates the los

s of some Bgl II restriction sites.

Fig. 1 Southern blot analysis of Meloidogyne exigua genomic DNA. For each restriction enzyme, 2µg of total genomic DNA were digested to completion, fractionated on a 1% agarose gel, transferred on to a nylon membrane and hybridized with the≈ 270 bp Bgl II fragments.

1993). In particular, such a feature may lead to the development of taxonomic markers suitable for molecular diagnostics, as has been shown in the case of nematodes of agronomic interest (reviewed in Grenier et al., 1997). In this paper, we report the identi cation, molecular cloning, genomic organization and sequence analysis of a new Bgl II satDNA from the coffee RKN M. exigua. We show that this sequence is species-speci c and can therefore be used as an identi cation tool for pest management programmes. Moreover, the results of this study are compared with other RKN satDNAs in order to discuss the general evolutionary processes acting on these sequences.

RESULTSDetection and cloning of a Bgl II satellite DNA in Meloidogyne exigua Total genomic DNA of M. exigua (isolate ex1) was digested to completion with a set of 11 restriction enzymes in order to detect highly repetitive sequences. With four enzymes (AluI, Bgl II, RsaIFig. 2 Time-course digestion of Meloidogyne exigua genomic DNA with Bgl II. For each lane, 2µg of total genomic DNA were digested, fractionated on a 1% agarose gel, transferred on to a nylon membrane and hybridized with the≈ 270 bp Bgl II fragments.

MOLECULAR PLANT PATHOLOGY (2002) 3(6), 431–437© 2002 BLACKWELL SCIENCE LTD

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