The genetic identity of alien chromosomes in potato breeding lines revealed by sequential GISH and FISH analyses using chromosome-specific cytogenetic DNA markers
Fg. Dong et al., The genetic identity of alien chromosomes in potato breeding lines revealed by sequential GISH and FISH analyses using chromosome-specific cytogenetic DNA markers, GENOME, 44(4), 2001, pp. 729-734
Genomic in situ hybridization (GISH) is one of the most popular and effecti
ve techniques for detecting alien chromatin introgressed into breeding line
s; however, GISH analysis alone does not reveal the genetic identity of the
alien chromosomes. We previously isolated a set of bacterial artificial ch
romosomes (BACs) specific to each of the 12 potato chromosomes. These BAC c
lones can be used as chromosome-specific cytogenetic DNA markers (CSCDMs) f
or potato chromosome identification. Here we demonstrate that GISH and fluo
rescence in situ hybridization (FISH), using CSCDMs, can be performed seque
ntially on the same chromosome preparations. Somatic metaphase chromosomes
prepared using an enzymatic digestion and "flame-drying" procedure allows r
epeated probing up to five times without significant damage to chromosome m
orphology. The sequential GISH and FISH analyses reveal the genomic origin
and genetic identity of the alien chromosomes in a single experiment and al
so determine whether an alien chromosome has been added to the genetic back
ground of potato or is substituting for a homoeologous potato chromosome. T
he sequential GISH and FISH procedures should be widely applicable for germ
plasm characterization, especially in plant species with small-sized chromo
somes.