D. Shibata et al., Establishment of framework P1 clones for map-based cloning and genome sequencing: direct RFLP mapping of large clones, GENE, 225(1-2), 1998, pp. 31-38
Large insert capacity, clone stability and convenient propagation in Escher
ichia coli have made bacterial artificial chromosome and phage P1 vector-ba
sed libraries the first choice for large-scale sequencing projects, and the
se libraries have also proven useful for chromosome walking. The applicatio
n of these libraries for either purpose is greatly facilitated by the estab
lishment of a set of framework clones distributed across the genome. Using
a Pi-based library of Arabidopsis thaliana with genomic inserts of 70-90 kb
(Liu, Y.-G., Mitsukawa, N., Vazquez-Tello, A., Whittler, R.F., 1995. Gener
ation of a high-quality P1 library of Arabidopsis suitable for chromosome w
alking. Plant J. 7, 351-358), we have now established such a set of framewo
rk clones. To date, such clones have usually been identified by hybridizati
on to smaller, previously mapped clones that detect restriction fragment le
ngth polymorphisms (RFLPs). In order to establish framework clones more eff
iciently, we refined protocols for PI clone DNA isolation and RFLP detectio
n in order to employ whole P1 clones directly as probes. This strategy enab
led a very high rate of RFLP detection, and obviated the need to screen the
P1 library with smaller RFLP probes. Altogether 95 clones were mapped prov
iding a framework into which further clones can be integrated by physical o
verlap. (C) 1998 Elsevier Science B.V. All rights reserved.