江苏省植物细胞遗传学研究回顾与展望

20世纪初“遗传的染色体学说”的提出和证明标志着细胞遗传学交叉学科建立,伴随相关学科的发展,20世纪60年代末期细胞遗传学又与分子遗传学相结合,建立发展了分子细胞遗传学交叉学科。分子细胞遗传学以DNA分子原位杂交技术为核心,不断拓展应用领域,为生命科学研究提供了直观、高效的技术手段。原位杂交技术与基因组、细胞生物学等技术结合,被广泛应用于人类、动物、植物的起源、进化、驯化等基础研究和远缘杂交、染色体工程等应用研究。通过形象地展示DNA、RNA、蛋白质在细胞中的实际位置,揭示DNA序列之间的实际位置和顺序、亲缘物种间的进化关系和结构重排、基因组拼接序列的质量、转录水平RNA和翻译水平蛋白质的位置和数量变化等。江苏省遗传学会会员单位南京农业大学、扬州大学、南京林业大学、江苏师范大学、徐淮地区农科院等自20世纪中期开展细胞遗传学理论技术研究,伴随学科发展不断创新,建立了较完善的分子细胞遗传技术体系,并成功应用于开展植物系统进化、远缘杂交、染色体工程、基因组学等研究,取得了一批研究成果。本文将主要综述江苏省在该领域取得的重要进展,并展望未来发展方向。

Pm21 from Haynaldia villosa Encodes a CC-NBS- LRR Protein Conferring Powdery Mildew Resistance in Wheat

Dear Editor Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici （Bgt）, is a destructive disease of wheat throughout the world. One of the most important environmental-friendly and economical methods to reduce wheat loss caused by Bgt is to develop highly resistant varieties （Kuraparthy et al., 2007）. Pm21 from the wild species Haynaldia villosa （also known as Dasypyrum villosum） confers high resistance to Bgt in wheat throughout all growth stages. It has now become one of the most highly effective genetic loci introgressed into wheat from wild species, and the commercial varieties harboring Pm21 have been widely used in wheat production with more than 4 million hectares in China.

Induction and transmission of wheat-Haynaldia villosa chromosomal translocations

In order to develop more wheat-Haynaldia villosa translocations involving different chromosomes and chromosome segments of H. villosa, T. durum-H, villosa amphiploid was irradiated with ^60Co γ-rays at doses of 800, 1,200, and 1,600 rad. Pollen collected from the spikes 1, 2, and 3 days after irradiation were transferred to emasculated spikes of the common wheat cv. ‘Chinese Spring＇. Genomic in situ hybridization was used to identify wheat-H, villosa chromosome translocations in the M1 generation. Transmission of the identified translocation chromosomes was analyzed in the BC1, BC2, and BC3 generations. The results indicated that all three irradiation doses were highly efficient for inducing wheat-alien translocations without affecting the viability of the M1 seeds. Within the range of 800-1,600 rad, both the efficiency of translocation induction and the frequency of interstitial chromosome breakage-fusion increased as the irradiation dosage increased. A higher translocation induction frequency was observed using pollen collected from the spikes 1 day after irradiation over that of 2 or 3 days after irradiation. More than 70% of the translocations detected in the M1 generation were transmitted to the BC1 through the female gametes. All translocations recovered in the BC1 generation were recovered in the following BC2, and BC3 generations. The transmission ability of different translocation types in different genetic backgrounds showed an order of ‘whole-arm translocation 〉 small alien segment translocation 〉 large alien segment translocation＇, through either male or female gametes, In general, the transmission ability through the female gametes was higher than that through the male gametes. By this approach, 14 translocation lines that involved different H. villosa chromosomes have been identified in the BC3 using EST-STS markers, and eight of them were homozygous.

Characterization of a Putative New Semi-Dominant Reduced Height Gene, Rht_NM9, in Wheat(Triticum aestivum L.)

Plant height is an important agronomic trait in cereal crops, and can affect both plant architecture and grain yield. New dwarfing genes are required for improving the genetic diversity of wheat. In this study, a novel dwarf mutant, NM9, was created by treating seeds of the wheat variety NAU9918 with ethyl methanesulfonate （EMS）. NM9 showed obvious phenotypic changes, which were distinct from those caused by other dwarfing genes, especially the reduced plant height, increased effective tiller number, and elongated spike and grain length. The reduced plant height in NM9 was attributable to a semi-dominant dwarfing gene Rht__NM9, which was flanked by two closely linked SNP markers, SNP34 and SNP41, covering an 8.86-Mb region on the chromosome arm 2AS. The results of gibberellic acid （GA） sensitivity evaluation, comparative genomics analysis and allelism test indicated that RhtNM9 was neither allelic to Rht7 and Rht21 nor homoe- oallelic to Rht8, so Rht_NM9 was proposed to be a new dwarfing locus on the homoeologous group 2 chromosomes of wheat. Rht_NM9 has a negative effect on plant height and positive effects on effective tiller number and grain size, thus, Rht_NM9 could be used for elucidating the mechanisms underlying plant architecture and grain development.

Cytogenetic and molecular identification of three Triticum aestivum-Leymus racemosus translocation addition lines

Chromosome 2C from Aegilops cylindrica has the ability to induce chromosome breakage in common wheat （Tritivum aestivum）. In the BC1F3 generation of the T. aestivum cv. Chinese Spring and a hybrid between T. aestivum-Leymus racemosus Lr.7 addition line and T. aestivum-Ae, cylindrica 2C addition line, three disomic translocation addition lines （2n = 44） were selected by mitotic chromosome C-banding and genomic in situ hybridization. We further characterized these T. aestivum-L, racemosus translocation addition lines, NAU636, NAU637 and NAU638, by chromosome C-banding, in situ hybridization using the A- and D-genome-specific bacterial artificial chromosome （BAC） clones 676D4 and 9M13; plasmids pAsl and pSc119.2, and 45S rDNA; as well as genomic DNA of L. racemosus as probes, in combination with double ditelosomic test cross and SSR marker analysis. The translocation chromosomes were designated as T3AS-Lr7S, T6BS-Lr7S, and T5DS-Lr7L. The translocation line T3AS-Lr7S was highly resistant to Fusarium head blight and will be useful germplasm for resistance breeding.

Competitive Expression of Endogenous Wheat CENH3 May Lead to Suppression of Alien ZmCENH3 in Transgenic Wheat × Maize Hybrids

Uniparental chromosome elimination in wheat × maize hybrid embryos is widely used in double haploid production of wheat. Several explanations have been proposed for this phenomenon, one of which is that the lack of cross-species CENH3 incorporation may act as a barrier to interspecies hybridization. However, it is unknown if this mechanism applies universally. To study the role of CENH3 in maize chromosome elimination of wheat x maize hybrid embryos,？cos, maize ZmCENH3 and wheat aTaCENH3-B driven by the constitutive CaMV35S promoter were transformed into wheat variety Yangmai 158. Five transgenic lines for ZmCENH3 and six transgenic lines for ctTaCENH3-B were identified. RT-PCP analysis showed that the transgene could be transcribed at a low level in all ZmCENH3 transgenic lines, whereas transcription of endogenous wheat CENH3 was significantly up-regulated. Interestingly, the expression levels of both wheat CENH3 and ZmCENH3 in the ZmCENH3 transgenic wheat × maize hybrid embryos were higher than those in the non-transformed Yangmai 158 × maize hybrid embryos. This indicates that the alien ZmCENH3 in wheat may induce competitive expression of endogenous wheat CENH3, leading to suppression of ZmCENH3 over-expression. Eliminations of maize chromosomes in hybrid embryos of ZmCENH3 transgenic wheat ×maize and Yangmai 158 x maize were compared by observations on micronuelei presence, by marker analysis using maize SSRs （simple sequence repeats）, and by FISH （fluorescence in situ hybridization） using 45S rDNA as a probe. The results indicate that maize chromosome elimination events in the two crosses are not sigmficantly different. Fusion protein ZmCENH3- YFP could not be detected in ZmCENH3 transgenic wheat by either Western blotting or immnunostaining, whereas accumulation and loading of the αTaCENH3-B-GFP fusion protein was normal in aTaCENH3-B transgenic lines. As ZmCENH3-YFP did not accumulate after AM114 treatment, we speculate that low levels of ZmCENH3 protein in transgenic wheat may be one of the factors that lead to failure of suppression of maize chromatin elimination in ZmCENH3 transgenic wheat × maize hybrids.