Characterization of a species-specific repetitive DNA from a highly endangered wild animal, Rhinoceros unicornis, and assessment of genetic polymorphism by microsatellite associated sequence amplification (MASA)

We have cloned and sequenced a 906 bp EcoRI repeat DNA fraction from Rhinoc eros unicornis genome. The contig pSS(R)2 is AT rich with 340 A (37.53%), 1 87 C (20.64%), 173 G (19.09%) and 206 T (22.74%). The sequence contains MAL T box, NF-E1, Poly-A signal, lariat consensus sequences, TATA box, translat ional initiation sequences and several stop codons. Translation of the cont ig showed seven different types of protein motifs, among which, EGF-like do main cysteine pattern signatures and Bowman-Birk serine protease inhibitor family signatures were prominent. The presence of eukaryotic transcriptiona l elements, protein signatures and analysis of subset sequences in the 5' r egion from 1 to 165 nt indicating coding potential (test code value = 0.97) suggest possible regulatory and/or functional role(s) of these sequences i n the rhino genome. Translation of the complementary strand from 906 to 706 nt and 190 to 2 nt showed proteins of more than 7 kDa rich in non-polar re sidues. This suggests that pSS(R)2 is either a part of, or adjacent to, a f unctional gene. The contig contains mostly non-consecutive simple repeat un its From 2 to 17 nt with varying frequencies, of which four base motifs wer e found to be predominant. Zoo-blot hybridization revealed that pSS(R)2 seq uences are unique to R. unicornis genome because they do not cross-hybridiz e, even with the genomic DNA of South African black rhino Diceros bicornis. Southern blot analysis of R unicornis genomic DNA with pSS(R)2 and other s ynthetic oligo probes revealed a high level of genetic homogeneity, which w as also substantiated by microsatellite associated sequence amplification ( MASA), Owing to its uniqueness, the pSS(R)2 probe has a potential applicati on in the area of conservation biology for unequivocal identification of ho rn or other body tissues of R unicornis. The evolutionary aspect of this re peat fraction in the context of comparative genome analysis is discussed. ( C) 1999 Elsevier Science B.V. All rights reserved.