Dissection of Alien Chromosomes
The Gc system can also be used to dissect alien (A) chromosomes introduced into common wheat. The cross scheme is simple: First, make a cross between the Gc disomic addition and an alien disomic addition; then, backcross the hybrid to the alien addition to produce plants disomic for the alien chromosome and monosomic for the Gc chromosome (42 +A” + Gc') (Endo 2007). In the progeny of this plant, we can find plants carrying structurally changed alien chromosomes, as well as aberrant wheat chromosomes.
By FISH/GISH we can undoubtedly identify deletions and translocations of alien chromosomes and can tell the exact breakpoint of a translocation between alien and wheat chromosomes (Fig. 8.4). The Gc system was successfully applied to produce dissection lines of various alien chromosomes in common wheat, such as barley chromosomes (2H, Joshi et al. 2011; 3H, Sakai et al. 2009; 4H, Sakata et al. 2010; 5H, Ashida et al.2007; 7H, Masoudi-Nejad et al. 2005), rye 1R chromosomes (Tsuchida et al. 2008; Gyawali et al. 2009, 2010) and rye B chromosomes (Endo et al. 2008). All possible kinds of rearrangements of alien chromosomes have been obtained (Fig. 8.5).
PCR-Based Mass Selection of Gc-Induced Deletions for Specific Chromosomes
The chromosomal deficiencies in the deletion stocks of common wheat were cytologically identified and characterized by chromosome banding. Then the breakpoints of the deleted chromosomes were analyzed with DNA markers. Once the
Fig. 8.4 A homozygous reciprocal translocation between barley chromosome 3H and a wheat chromosome
Fig. 8.5 Examples of alien chromosome aberrations induced by the Gc system. The green FISH signals indicate the subtelomeric repeats HvT01 for the barley 5H and 7H chromosomes derived from a cultivar Betzes, the pSc200 repeats for the rye 1R chromosome from a cultivar Imperial, and the E1100 repeats for the rye B chromosome from a Siberian cultivar. The GISH signals are shown in red for all chromosomes
Fig. 8.6 A schematic diagram showing the PCR-based mass selection
positions of the DNA markers are located on a chromosome, more deficiencies of the chromosome can be identified using chromosome-specific DNA markers and appropriate aneuploids. For example, using PCR-based 6B-specific markers, a search was made for deficiencies of chromosome 6B among the progeny from a cross between nullisomic 6B-tetrasomic 6A and a monosomic 2C addition line as the pollen parent (Fig. 8.6). Any deficiencies occurring in the 6B chromosome within the two markers could be detected by PCR because no 6B homologue was transmitted from the female parent. Thus, 102 (5.0 %) of the 2,041 hybrid plants were found to have a deficiency in either or both chromosome arms (unpublished data). In a similar way, Gc-induced deletions of alien chromosomes in common wheat can be identified by PCR. This PCR selection should find deletions overlooked by cytology and moreover probably detect structural changes that do not exist in the root tips but exist in aerial parts of plants. Joshi et al. (2013) conducted PCR analysis in 81 plants carrying a cytologically normal-appearing 2H chromosome in root tips and detected 2H aberration in the leaves of 6 of them. This fact implied the ongoing production of aberrations after fertilization.
Thanks to the next-generation sequencing technologies, sequencing the entire genomes of wheat, barley and rye has become a reality. Several chromosomal landmarks will be needed to assemble contigs into supercontigs or even into chromosomes. The Gc system will help provide a virtually limitless number of such landmarks, namely breakpoints of deleted or translocated chromosomes of the new aneuploids of common wheat.
Lastly, I wish to express my gratitude to Prof. Bikram S. Gill for his encouragement to establish deletion stocks common wheat by using them for his wheat chromosome mapping studies, otherwise I would not have done these works mentioned above.