Presence of a chloroplast DNA sequence in an autonomous circular DNA molecule in cultured rice cells (Oryza sativa L.)

• Published in: Plant and cell physiology 1995-06, Vol.36 (4), p.717-725
• Main Authors: Cuzzoni, E. (Universita di Pavia (Italia)), Giordani, C, Stampacchia, O, Bolchi, A, Malcevschi, A, Ottonello, S, Ferretti, L, Sala, F
• Format: Article
• Language: English
• Subjects: ADN; BIOLOGIA MOLECULAR; Biological and medical sciences; BIOLOGIE MOLECULAIRE; CELL CULTURE; Cells, Cultured; Chloroplast DNA; CHLOROPLASTE; CHLOROPLASTS; Classical genetics, quantitative genetics, hybrids; CLOROPLASTO; CULTIVO DE CELULAS; CULTURE DE CELLULE; DNA; DNA amplification; DNA Probes; DNA, Chloroplast - analysis; DNA, Circular - analysis; Extrachromosomal DNA; Fundamental and applied biological sciences. Psychology; Genetics of eukaryotes. Biological and molecular evolution; MOLECULAR BIOLOGY; NUCLEOTIDE SEQUENCE; Oryza - cytology; Oryza - genetics; ORYZA SATIVA; Polymerase Chain Reaction; Pteridophyta, spermatophyta; Pulsed-field gel electrophoresis; SECUENCIA NUCLEICA; Sequence Analysis, DNA; SEQUENCE NUCLEIQUE; Vegetals
• ISSN: 0032-0781; 1471-9053; 1471-9053
• DOI: 10.1093/oxfordjournals.pcp.a078813

Sequence analysis of twelve DNA fragments, which had previously been found to be extensively amplified in suspension-cultured rice cells, revealed that two of them, isolated on plasmids designated pE10 and pE11, have sequences identical to distinct regions of chloroplast DNA (ct-DNA). Both sequences are part of an extrachromosomal circular DNA molecule (ECD). The molecular structure of the ECD was investigated by a combination of restriction analysis, standard and pulsed-field gel electrophoresis, hybridization with ct-DNA probes and amplification by the polymerase chain reaction in the presence of oligonucleotide primers homologous to selected regions of rice ct-DNA. The results showed that a continuous and unrearranged stretch of ct-DNA from the long single-copy region, of at least 28 kbp in length, is present in the ECD. It was estimated that the number of copies of the ECD in cultured cells was almost equivalent to that of ct-DNA molecules in rice leaves, while the ratio of ECD to ct-DNA molecules in the cultured cells was approximately 200: 1.