Start Sites of Bidirectional DNA Synthesis at the Human Lamin B2 Origin
Gulnara Abdurashidova,
1*
Marta Deganuto,
1*
Raffaella Klima,
1
Silvano Riva,
2
Giuseppe Biamonti,
2
Mauro Giacca,
3
Arturo Falaschi
1
The initiation sites of bidirectional synthesis at the DNA
replication origin located at the 3' end of the human lamin B2 gene
were investigated. RNA-primed nascent DNA molecules were subjected to
second-strand synthesis with appropriate primers, amplified by
ligation-mediated polymerase chain reaction, and size fractionated.
Evidence for precise start sites was obtained. Exploration of close to
1 kilobase, coupled to inhibition of Okazaki fragment synthesis,
demonstrates that the leading strands initiate at precise nucleotides
on either helix, overlapping by three base pairs, within the area bound
to a protein complex possibly analogous to the prereplicative complex
of yeast.
1 Molecular Biology and
3 Molecular Medicine Units, International Centre for
Genetic Engineering and Biotechnology, Padriciano 99, 34012 Trieste,
Italy.
2 Istituto di Genetica Biochimica ed Evoluzionistica
of the Consiglio Nazionale delle Ricerche, Via Abbiategrasso 207, 27100 Pavia, Italy.
*
These authors contributed equally to this work.
To whom correspondence should be addressed. E-mail:
falaschi{at}icgeb.trieste.it
