Different DNA methylation pattern in A and B chromosomes of Crepis capillaris detected by in situ nick-translation. Comparison with molecular methods.
Experiments were performed on Crepis capillaris callus lines with 0, 1 and 2 B chromosomes and on hairy root lines without or with 1 and 2 B chromosomes. Comparison of HPLC results for DNA from calli differing in number of B chromosomes did not reveal any significant differences in methylation level (30.4 +/- 1.1%, 30.9 +/- 1.2%, 31.7 +/- 1.7% in lines without or with one or two B chromosomes respectively) which could be attributed to the number of B chromosomes. Restriction patterns obtained after DNA digestion with HhaI, HpaII, MspI or HaeIII (i.e. restriction enzymes sensitive to cytosine methylation) were similar in calli and apical root segments and also did not depend on the presence or number of B chromosomes. Methylation of B chromosomes higher than that of A chromosomes was demonstrated by fluorescent in situ nick translation driven by HpaII, MspI or HaeIII in metaphase chromosomes. After short digestion (I and 3 h), B chromosomes, in contrast to A chromosomes, were weakly labelled or not labelled at all, which indicates longer distances between target sequences containing unmethylated cytosine in the former.
Luchniak
Piotr
P
Department of Cytogenetics and Plant Molecular Biology, University of Lódź, Poland.
Maluszynska
Jolanta
J
Olszewska
Maria J
MJ
D002875:Chromosomes / Q000235:genetics* / Q000648:ultrastructure; D027468:Crepis / Q000235:genetics*; D019175:DNA Methylation; D018744:DNA, Plant / Q000235:genetics* / Q000302:isolation & purification; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005821:Genetic Techniques; D008677:Metaphase; D018517:Plant Roots; D014176:Protein Biosynthesis; D015183:Restriction Mapping; D012231:Rhizobium / Q000235:genetics; D013050:Spectrometry, Fluorescence
D018744:DNA, Plant