Cytogenetic And Molecular Research Of The Germplasm From Octoploid Tritileymus And Triticum Durum

Leymus mollis (Trin.) Pilger (NsNsXmXm,2n=28), a wild relative of common wheat,possesses many potentially valuable traits that could be transferred to common wheat, such asstrong stems, large spikes, multiple spikelets, tolerance of drought, cold, salts and infertilesoils, and resistance to fungal and bacterial diseases. L. mollis is an allotetraploid species withan Ns genome originated from the genus Psathyrostachys Keng, and an Xm genome ofunknown origin. Octoploid Tritileymus, obtained from the hybridization of Triticum aestivumand L. mollis, is a novel synthetic germplasm with many extreme agronomic characteristicsfrom L. mollis, such as long spikes, large seeds, strong stem, tolerance to cold and drought,resistance to several fungal an bacterial diseases, although with wrinkled kernels, low self-fertility and late maturing. In order to further exploit and utilize the excellent germplasmresource of L. mollis, the cross between octoploid Tritileymus M842-16(AABBDDNsNs,2n=56) and Triticum durum cv. D4286(AABB,2n=28) was made, and a series of progeniesare selected and identified cytogenetic methods such as in situ hybridization and DNAmolecular markers. The main results as followed:(1) Molecular cytogenetic identification of progenies from the cross of octoploidTritileymus M842-16and T. durum D4286: nine types of Tritileymus substitution lines, withagronomic uniformity and genetic stability, are identified from the F5-F7progenies ofoctoploid Tritileymus M842-16and Triticum durum cv. D4286by cytogenetic, GISH(Genomic in situ hybridization) and EST-STS (Expressed sequence tag-sequence tagged sites)methods. Two types contain two Ns chromosomes, and the alien chromosomes are2Ns and3Ns, respectively. Three types contain four Ns chromosomes, and the alien chromosomecompositions are2Ns/3Ns,3Ns/6Ns, and3Ns/7Ns, respectively. Four types contain six Nschromosomes, and the alien chromosomes compositions are2Ns/3Ns/7Ns,2Ns/6Ns/7Ns,3Ns/5Ns/7Ns and3Ns/6Ns/7Ns, respectively. There are more than twenty lines with Nschromosomes still need study.(2) Molecular cytogenetic identification of diploid substitution line DM57: A wheat-L. mollis diploid3D/3Ns substitution line DM57, is described by GISH, FISH (Fluorescent insitu hybridization), SSR (simple sequence repeat), and EST-STS methods. Mitotic andmeiotic GISH using the genomic DNA of P. huashanic as a probe shows that, two Nschromosomes are detected in the root tip cells, and could pair completely to form a ringbivalent in the wheat background. The FISH, SSR and EST-STS show that,3D chromosomesare absent and replaced by3Ns chromosomes from octoploid Tritileymus. So, DM57is a3D/3Ns Tritileymus substitution line with40chromosomes from wheat and two3Nschromosomes from L. mollis. Compared to common wheat parent7182, DM57show moreand longer spikes, heavier1,000-kernal weight, and excellent resistance to leaf rust, but withhigher plant height and lower self-fertility. A hybridization pattern of repeated sequence pAs1on the3Ns chromosomes of DM57is observed, and could be used to identify3Nschromosome of L. mollis. Compared to the hybridization pattern of3D chromosomes inwheat,3Ns show the similar pattern on the long-and short-arm termini, and on thesubterminal long arms, respectively, but no hybridization site on the middle part of short-arm.And this pattern could be used as the specific landmark to identify3Ns in the progenieswheat-L. mollis.(3) Molecular cytogenetic identification of multiple substitution line DM50: A multiplesubstitution line DM50is described by GISH, SSR and EST-STS methods. Mitotic andmeiotic GISH using the genomic DNA of P. huashanic as a probe shows that, DM50is acytogenetic stable line with thirty-six chromosomes from wheat and six Ns chromosomesfrom L. mollis. SSR analysis show that,3D,6D and7D chromosomes are absent in DM50.EST-STS analysis shows that, the chromatin of3Ns,6Ns and7Ns from L. mollis are detected.Compared to common wheat parent7182, the spikes per plants, spike length,1,000-kenelweight of DM57and DM50are improved significantly, but the self-fertility of them isdeclined.(4) FISH analysis of four multiple substitution lines: The chromosome composition offour multiple substitution lines are identified by GISH and FISH. The results show that,DM92, with four Ns chromosomes, lose the3D and6D chromosomes. DM96, with four Nschromosomes, lose the2D,3D and6D chromosomes. DM99, with six Ns chromosomes, lostthe2D,3D and7D chromosomes. DM108, with six Ns chromosomes, lose the3D,5D and7D chromosomes. And the affiliated groups are of one-to-one correspondence between thelost chromosomes and alien chromosomes.(5) Screening of special EST-STS primers to Ns chromosomes from L. mollis: Thirty- four out of five hundred and fifty-nine EST-STS primers, that show polymorphism betweenNs genome of L. mollis and A, B, D genome of wheat, are selected to detect the Ns chromatinof the progenies of octoploid Tritileymus×Triticum durum D4286. Seven EST-STS primersare from group1, four from group2, four from group3, two from group4, two from group5,six from group6, and nine from group7.