Vrána, J., Kubaláková, M., Číhalíková, J., Valárik, M., Doležel, J.
BIOLOGIA PLANTARUM
59:
445-455,
2015
Keywords:
chromosome sorting, complexity reduction, genome analysis
Abstract:
Flow-sorted chromosomes have been used to simplify analyses of complex plant genomes. In bread wheat, majority of
studies involve cultivar Chinese Spring, a genotype chosen for sequencing. Telosomic lines developed from this cultivar
enable isolation by flow sorting chromosome arms, which represent less than 3.4 % of the genome. However, access to
other wheat cultivars is needed to allow mapping and cloning useful genes. In these cultivars, cytogenetic stocks are not
readily available and only one chromosome (3B) can be sorted. Remaining chromosomes form composite peaks on flow
karyotypes and cannot be sorted. In order to overcome this difficulty, we tested a pragmatic approach in which
composite chromosome peaks are dissected to smaller sections. The analysis of chromosome composition in sorted
fractions confirmed feasibility of obtaining sub-genomic fractions comprising only a few chromosomes. Usually one of
the chromosomes was more abundant and the frequencies of dominant chromosomes in sorted fractions ranged from
16 % (chromosome 7B) to 80 % (chromosome 2B). The enrichment factor, calculated as the relative proportion of
chromosomal DNA in the wheat genome to the proportion of chromosomal DNA in a sorted fraction, ranged from
3.2-fold (7B) to 16.4-fold (5D). At least a 5-fold enrichment can be obtained for 17 out of 21 wheat chromosomes.
Moreover, we show that 15 out of the 21 chromosomes can be sorted without being contaminated by their homoeologs.
These observations provide opportunities for constructing sub-genomic large-insert DNA libraries, optical mapping,
and targeted sequencing selected genome regions in various cultivars of wheat. The availability of fractions enriched for
chromosomes of interest and free of contaminating homoeologs will increase the efficiency of research projects and
reduce their costs as compared to whole genome approaches. The same methodology should be feasible in other plants
where single chromosome types cannot be sorted.
Fulltext: contact IEB authors
IEB authors: Jarmila Číhalíková,
Jaroslav Doležel,
Marie Kubaláková,
Miroslav Valárik,
Jan Vrána
