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Local repeat sequence organization of an intergenic spacer in the chloroplast genome ofChlamydomonas reinhardtii leads to DNA expansion and sequence scrambling: A complex mode of “copychoice replication”?

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Abstract

Parent-specific, randomly amplified polymorphic DNA (RAPD) markers were obtained from total genomic DNA ofChlamydomonas reinhardtii. Such parent-specific RAPD bands (genomic fingerprints) segregated uniparentally (through mt+) in a cross between a pair of polymorphic interfertile strains ofChlamydomonas (C. reinhardtii andC. minnesotti), suggesting that they originated from the chloroplast genome. Southern analysis mapped the RAPD-markers to the chloroplast genome. One of the RAPD-markers, “P2” (1.6 kb) was cloned, sequenced and was fine mapped to the 3 kb region encompassing 3′ end of 23S, full 5S and intergenic region between 5S and psbA. This region seems divergent enough between the two parents, such that a specific PCR designed for a parental specific chloroplast sequence within this region, amplified a marker in that parent only and not in the other, indicating the utility of RAPD-scan for locating the genomic regions of sequence divergence. Remarkably, the RAPD-product, “P2” seems to have originated from a PCR-amplification of a much smaller (about 600 bp), but highly repeat-rich (direct and inverted) domain of the 3 kb region in a manner that yielded no linear sequence alignment with its own template sequence. The amplification yielded the same uniquely “sequence-scrambled” product, whether the template used for PCR was total cellular DNA, chloroplast DNA or a plasmid clone DNA corresponding to that region. The PCR product, a "unique" new sequence, had lost the repetitive organization of the template genome where it had originated from and perhaps represented a “complex path” of copy-choice replication.

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Author notes

  1. Mahendra D. Wagle

    Present address: Institute of Molecular Agrobiology, 1, Research Link, National University of Singapore, 117 604, Singapore

Authors and Affiliations

  1. Department of Biological Sciences, Tata Institute of Fundamental Research, Homi Bhabha Road, Colaba, 400 005, Mumbai, India

    Mahendra D. Wagle, Subhojit Sen & Basuthkar J. Rao

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  1. Mahendra D. Wagle
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  2. Subhojit Sen
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  3. Basuthkar J. Rao
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Correspondence to Basuthkar J. Rao.

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Wagle, M.D., Sen, S. & Rao, B.J. Local repeat sequence organization of an intergenic spacer in the chloroplast genome ofChlamydomonas reinhardtii leads to DNA expansion and sequence scrambling: A complex mode of “copychoice replication”?. J. Biosci. 26, 583–594 (2001). https://doi.org/10.1007/BF02704757

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