Location: Forage and Range ResearchTitle: Microdissection and chromosome painting of the alien chromosome in an addition line of wheat-Thinopyrum intermedium Author
Submitted to: PLoS One
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/11/2013
Publication Date: 8/14/2013
Citation: Deng, C.L., Bai, L.L., Fu, S.L., Yin, W.B., Zhang, Y.X., Chen, Y.H., Wang, R., Zhang, X.Q., Han, F.P., Hu, Z.M. 2013. Microdissection and chromosome painting of the alien chromosome in an addition line of wheat-Thinopyrum intermedium. PLoS One. 8(8):e72564. DOI:10.1371/journal.pone.0072564. Interpretive Summary: Intermedium wheatgrass is an important forage grass as well as a valuable gene reservoir for wheat improvement. The genetic makeup of intermediate wheatgrass is so complicated that different interpretations and conclusions have been put forth in scientific literature. It is generally accepted that intermediate wheatgrass is an autoallohexaploid species. A lot of hybrid derivatives of wheat crossed with intermediate wheatgrass are available, some of which offer an opportunity to investigate the grass chromosomes individually as to their genomic origin and genetic markers for important agronomic traits that they are carrying. We studied the alien chromosomes in an addition line Tai-27 by microdissection and determined them to be two pairs of chromosomes belonging to St genome by genomic in situ hybridization. The genome constitution of intermediate wheatgrass was also elucidated and revealed by multi-color chromosome painting. A repetitive sequence specific to H- and V-genome can be used to distinguish the Js genome from J and St. These techniques are useful in research aiming to characterize and map individual chromosomes in intermediate wheatgrass or other polyploid species.
Technical Abstract: The chromosome painting is an efficient tool for chromosome research. However, plant chromosome painting is relatively underdeveloped. In this study, chromosome painting was developed and used to identify alien chromosomes in TAi-27, a wheat-Thinopyrum intermedium addition line, and chromosomes of the three different genomes of Th. intermedium. The smallest alien chromosome was microdissected from root-tip metaphase chromosomes of TAi-27 with a fine glass needle. DNA was amplified by DOP-PCR (Degenerate oligonucleotide primed - polymerase chain reaction) from the isolated chromosome. The amplification products were used as the probe to hybridize with metaphase chromosomes of TAi-27 and Th. intermedium. Results showed that hybridization signals were observed in all regions of a pair of smallest alien chromosomes and pericentromeric area of another pair of alien chromosomes in TAi-27, indicating that the probe from microdissected chromosome is species specific. In Th. intermedium, wide and strong hybridization signals were distributed mainly on pericentromeric area of 14 chromosomes, narrowly on centromeric region of another 9 chromosomes, and none or very weak on the remaining chromosomes. The sequential FISH/GISH on Th. intermedium chromosomes using amplified DNA from the microdissected alien chromosome and the genomic DNA of Pseudoroegneria spicata (St genome) indicated that the microdisseted chromosome belonged to St genome. Sequential FISH on Th. intermedium chromosomes using amplified DNA from the microdissected alien chromosome and pDbH12 (Js genome specific probe) showed that three genomes (Js, J and St) in Th. intermedium could be distinguished. Our results indicated that the two pairs of alien chromosomes in Tai-27 should be from St genome, of which the smallest alien chromosomes should be trucated ones. Our results suggested that chromosome painting could be feasible in some plants, such as some progenies from wide crosses and polyploid plants and can be used to detect alien chromosome variation. The repetitive sequences distribution may be similar in different chromosomes within St genome, but different within Js genome, different between non-homologous chromosomes, and different among different genomes in Th. intermedium. This information promotes the usage of chromosome painting technique in plant research area and helpful for understanding the evolution of different genomes in polyploid plants, such as Th. intermedium.