Chromosome Evolutionary Analysis Of Wheat Using Oligonucleotide Probe Multiplex FISH

Fluoresence in situ hybridization(FISH)is a powerful tool for studying chromosome variation.FISH using oligonucleotide probe multiplex#4(ONPM#4)developed by our laboratory and A genome oligonucleotide probes developed in this study was used to analyze chromosomal evolution of synthetic wheat,wheat ancestors and wheat-Thinopyrum bessarabicum chromosome translocations.The main results are as follows:Firstly,six oligonucleotide probes were designed by sequencing and bioinformatics analysis of Triticum monococcum genome.By FISH,significant variations of the signal number and distribution positions in different species were observed.The 45S rDNA sequence of wheat was dissected into 9 oligonucleotide probes and formed a multiplex probe for distinguishing the satellite chromosomes of wheat and Aegilops tauschii.Additionally,a tandem repeat oligonucleotide probe BS-HC CL135-2 which was newly developed using Thinopyrum bessarabicum genomic sequences was combined with the pre-developed probes for corresponding polymorphic chromosomes of different Aegilops tauschii accessions.Secondly,the chromosome composition of 13 Synthetic wheat(SW)lines derived from durum wheat Langdon and different Aegilops tauschii was analyzed by ONPM#4.It was found that there were abundant polymorphisms between the chromosomes of different D genome sources.A total of 30 polymorphic types were found including 5 for each of 2D,4D and 6D,6 for 5D,4 for 1D,3 for 3D,and 2 for 7D.Further analysis of 301 individual F8 plants from 13 SW lines(5 to 40 individuals for each line)showed that 49.17%of plants involved chromosomal variations,but the frequency varied in different lines,ranging from 8.33%to 100%.According to the analysis of variant chromosomes,the variation frequency of B genome is the highest(32.89%),followed by D(16.28%)and A(15.95%);For 21 pairs of chromosomes,20 pairs involved variations,of which 1B had the highest frequency of variation(18.27%),followed by 4B(6.64%),5A(5.98%),1D=4D(5.65%),4A(3.65%),1A(3.32%),7B(2.66%),3D(1.99%),5B=6B=2D(1.66%),2A=6A=2B=3B=6D(1.00%),3A(0.66%),7A(0.33%),5D(0.33%)and no significant variation in 7D was found.Thirdly,using ONPM#4,12 wild species of wheat and 2 cultivars was analyzed,obvious variations of repeat sequences were found in both A and B genomes in tetraploid wheat(AABB)in comparison with the original A and S genomes.However,in the second distant hybridization between AABB and DD,no obvious changes in A and B genome chromosome were found in comparison with the D genome chromosomes of Aegilops tauschii and AABB of tetraploid wheat.Finally,using ONPM#4 FISH combined with GISH(Genomic in situ hybridization),Chinese Spring-Thinopyrum bessarabicum translocation T2JS-2BS·2BL was transferred into three different cultivars Yangmai 6,Yannong 19 and Aisu 3 by backcrossing.Nine wheat lines with promise agronomic traits were obtained and identified.The introduction of T2JS-2BS·2BL significantly increased plant height,spike length and preharvest sprouting resistance.The average height of Aisu 3 was increased by 16.45 cm and 1000 kernel weight by 9.84 g,which provided new germplasm for breeding wheat with reduced plant height,more spikes and higher preharvest sprouting resistance.