Plant Genome I Conference
Town & Country Conference Center, San Diego, CA, November, 1992.
PG-I: CONSTRUCTION OF RICE GENOMIC LIBRARIES IN YEAST ARTIFICIAL
CHROMOSOMES FOR PHYSICAL MAPPING
CONSTRUCTION OF RICE GENOMIC LIBRARIES IN YEAST ARTIFICIAL
CHROMOSOMES FOR PHYSICAL MAPPING.
Yosuke Umehara, Akiko Miyazaki, Hiroshi Tanoue, Yuji Yasukochi,
Shoko Saji, Yoshiaki Otsuki*, Tatsuhiko Fujimura, Nori Kurata and
Yuzo Minobe, Rice Genome Research Program, National Institute of
Agrobiological Resources / Society for Techno-innovation of
Agriculture, Forestry and Fisheries, and *National Agriculture
Research Center, Kannondai 2-1-2, Tsukuba, 305 lbaraki, Japan.
(Fax +81-298-38-7468)
Use of yeast artificial chromosomes is a powerful too] for
analysis of complex genomes, due to the capability of cloning
large DNA fragments (50kb - 1Mb). For preparing the physical map
of rice genome, we constructed genomic libraries in yeast
artificial chromosomes. High molecular weight DNA was extracted
from liquid-cultivated cells of japonica rice (cv. Nipponbare)
embedded in agarose plugs, and digested partially with EcoRl or
with Notl. Digested DNA was fractionated by pulsed-field gel
electrophoresis and ligated with the YAC vector pYAC4 (for EcoRl
digest) or pYAC55 (for NotI digest). Ligated products were
fractionated again and used to transform yeast (Saccharomyces
cerevisiae ABI380). We have obtained 8,000 YAC clones with EcoRl
digests and 4,000 YAC clones with Notl digests. The average size
of inserts is about 350kb and the maximum size is more than 1Mb,
as revealed by the analysis of 240 randomly selected clones.
These libraries cover the rice genome about 5 times. Isolation
of YAC clones containing a fragment of the rice chromosome 6 is
now in progress using chromosome-specific RFLP and STS markers.
We have already begun chromosome walking from these marker points
on chromosome 6.
Return to Previous Page or Intl-PAG Homepage