| The development and application of oligonucleotide probes have greatly facilitated the development of chromosome identification and chromosome engineering and have been successfully applied in many species.Rye and Haynaldia villosa are important relatives of wheat which are widely used in wheat breeding.The development of oligonucleotide probe multiplexes that can simultaneously distinguish wheat-rye or wheat-Haynaldia villosa chromosomes can greatly improve the efficiency of chromosome engineering of wheat.New advances have been obtained as follows:Firstly,a new oligonucleotide probe multiplex named ONPM#5 containing pAsl-1,pAs1-3,pAs1-4,pAs1-6,AFA-3,AFA-4,pSc119.2-1,(AAC)10 and oligo-6 has been developed.After ONPM#5 FISH,high resolution karyotypes of Chinese Spring,Huixianhong,Jingzhouheimai and Huixianhong-Jingzhouheimai amphiploid Jinghui 1 were developed which can clearly identify all wheat and rye chromosomes after one round FISH.79 hybrids of Huixianhong/Jingzhouheimai and 12 of T.durum-Haynaldia villosa amphiploid/Jinsong 49(a hexaploid triticale)were analyzed.42 plants were found involving chromosome variations of wheat and rye,which included 10 plants with chromosome additions involving 1R,2R,4R and 5R,8 plants with translocations involving 1R and 2R,6 plants with perInv6B,and one 2DS trisomic telosomes,2 telosomes involving 6BS and 2DS,and 15 plants with complex translocations.Among them seven were confirmed by genomic in situ hybridization(GISH),which proved the potential of the probe multiplexes.The ONPM#4(Du et al.2017)was previously developed which contained pAs1-1,pAs1-3,pAs1-4,pAs1-6,AFA-3,AFA-4,pSc119.2-1 and(GAA)10 which not only clearly distinguished all wheat chromosomes but also enabled clear distinguishing of all Haynaldia villosa chromosomes after one round FISH.Reference karyotypes of Haynaldia villosa,T.durum and T.durum-Haynaldia villosa amphiploid were thus developed and used for wheat-Haynaldia villosa chromosome engineering.Based on ONPM#4 FISH,36 T.durum-Haynaldia villosa alien chromosome lines previously developed or introduced by Nanjing Agricultural University were analyzed in combined with GISH and molecular marker analysis.Among them,six T6VS·6AL translocations,4 wheat-Haynaldia villosa disomic additions(DA1V,DA2V,DA4V and DA6V),one monosomic telocentric addition(MtA2VL),one translocation T5VS·7DL,3 wheat-Haynaldia villosa substitutions DS3V(3D),DS6V(6A)and DS2V(2D),and one deletion with disomic additions DSde16VL(6A)were validated;Additionally,two reciprocal translocations including T5AS·3BS/T5AL·3BL and T6DS·7BL/T7BS·6DL were identified,the former translocation were observed in 92R178 and 5 cultivars,and the latter was only observed in 08R62-11(DSde16VL(6A)).Since the release of series of wheat-Haynaldia villosa translocations T6VS·6AL,which have become a key powdery mildew resistance parent and got widely used in wheat breeding in China since 1990s.To date,more than 30 cultivars have been released and planted in Hebei,Jiangsu,Sichuan,Gansu and Anhui provinces of China.In this study,20 cultivars derived from T6VS"6AL were validated using oligopainting,GISH and molecular marker analysis.14(70.0%)cultivars with T6VS·6AL and 6 without were observed.Among 14 cultivars with T6VS·6AL,5 also contained translocations T5AS·3BS/T5AL·3BL,2 contained perInv6B,2 contained T1RS·1BL,and the remaining 5 only had T6VS·6AL;for the other 6 cultivars,one with perInv6B,two with T1RS·1BL,and the other three showed regular karyotypes.Additionally,among 20 cultivars,total 53 polymorphic chromosomes of wheat were identified,where B-genome chromosomes had most polymorphisms(26 types),followed by A-genome(16 types),and D-genome chromosomes(11 types).The frqeuncy of each polymorphic chromosome varied from 2.4%to 100%.16 chromosomes inlcuding 1A-1,2A-2,3A-1,4A-1,7A-2,1B-1,4B-2,5B-1,6B-1,1D-1,2D-1,3D-1,4D-2,5D-2,6D-1 and 7D-1 showed a frequency more than 50%,indicating dominant types in the present cultivars. |
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