反义RNA技术在植物基因工程领域中的应用

反义RNA技术在植物基因工程领域中的应用包括：a．番茄和其他水果的成熟控制;b．植物的抗病性;c．改变花卉的颜色;d．植物淀粉合成的控制;e．油料植物种子中脂肪酸合成的控制;f．杂交种子生产中雄性不育性的控制;g．其他.     

Inheritance of spontaneous male sterility in peas

A plant with a mutant phenotype was observed in a Longittee cultivar. The plant was late in maturity, had white-translucent anthers, and was male sterile. The inheritance of this mutant was studied in a cross involving the mutant and the mother parent and their F₁, F₂, F₃ and BC₁F₁ generations. The results suggested that the sterile character was genetic and due to a recessive gene.     

URF13, a ligand-gated,pore-forming receptor for T-toxin in the inner membrane ofcms-T mitochondria

URF13 is the product of a mitochondrial-encoded gene (T-urfl3) found only in maize plants containing the Texas male-sterile cytoplasm (cms-T), and it is thought to be responsible for both cytoplasmic male sterility and the susceptibility ofcms-T maize to the fungal pathogensBipolaris maydis race T andPhyllosticta maydis. Mitochondria isolated fromcms-T maize are uniquely sensitive to pathotoxins (T-toxin) produced by these fungi and to methomyl (a commercial insecticide). URF13 acts as a receptor that specifically binds T-toxin to produce hydrophilic pores in the inner mitochondrial membrane. When expressed inEscherichia coli cells, URF13 also forms hydrophilic pores in the plasma membrane if exposed to T-toxin or methomyl. Topological studies established that URF13 contains three membrane-spanning -helices, two of which are amphipathic and can contribute to pore formation. Chemical crosslinking of URF13 was used to demonstrate the existence of URF13 oligomers incms-T mitochondria andE. coli cells. The ability of the carboxylate-specific reagent,N,N-dicyclohexycarbodiimide, to cross-link URF13 was used in conjunction with site-directed mutagenesis to establish that the URF13 tetramer has a central core consisting of a four--helical bundle which undergoes a conformational change after interaction with T-toxin or methomyl. Overall, the experimental evidence indicates that URF13 functions as a ligand-gated, pore-forming T-toxin receptor incms-T mitochondria.     

Expression of the CMS-associated urfS sequence in transgenic petunia and tobacco

The expression of a 25 kDa protein, encoded by the fused mitochondrial pcf gene, is associated with cytoplasmic male sterility (CMS) in petunia. To investigate the role of the 25 kDa protein in CMS we have transformed petunia and tobacco plants with constructs expressing a portion of the urfS sequence of the pcf cDNA which encodes the 25 kDa protein. The urfS sequence was fused with two different mitochondrial targeting sequences. The chimeric gene coding region was placed under the control of the CaMV 35S promoter or a tapetum-specific promoter. Expression of the PCF protein was obtained in mitochondria of transgenic petunia and tobacco plants, yet fertility of the plants was not affected. Analysis of the location of the urfS-encoded protein revealed that it fractionates primarily into the soluble fraction in the transgenic plants whereas the genuine 25 kDa protein is found primarily in the soluble fraction but also in the membrane portion of immature buds from CMS petunia plants. Fertile transgenic plants were obtained which expressed the 25 kDa protein in the tapetal layer of post-meiotic anthers, while CMS plants express the endogenous 25 kDa protein in both the tapetal layer and sporogenous tissue of pre-meiotic anthers.     

小麦提型不育系和杂交种种子生活力遗传改良初探

本试验共选用了4个提型不育系(A系)及其保持系(B系)、4个恢复系(R系)及其川7B/R4份F~1|代材料,用不同遗传背景的B、R系及川7B/R材料与A系杂交,种子成熟时收获干燥考种,度过休眠期后进行发芽试验。结果表明,父本对F~0|种子千粒重存在胚乳直感现象;川3A、川4A与其它B系杂交,其F~0|种子的千粒重、饱满度、发芽率和发芽势均有不同程度的提高;川4A×R和川6A×R的杂种种子千粒重多分别比川4A×川7B/R和川6A×川7B/R的高,但前种组合(A×R)的种子发芽和发芽率远不如后一种组合(A×川7B /R)的种子,且前种组合的穗发芽率也较高。作者认为,利用A系与农艺性状相近、但遗传背景各异的B系杂交,或在R系中输入抗提型细胞质负影响的高种子生活力基因, 是提高A系和杂交种种子生活力的值得注意的途径。 Abstract:The objective of this paper is trying to grope for ways of improving sced viability of A-line and hybrid in wheat with T.timopheevi cytoplasm.Four A lines and their B lines,4 restorers (R line) and 4 crosses of Chuan 7B/R were used.The combinations of A×B,A×R and A×Chuan 7B/R were madc,and 1 000-grain weight (GW),rate of pre-harvest sprouting(RPHS),germinating energy(GE) and germination percentage (GP) of their F0 seeds were investigated.The results showed that the GW,full weight,GE and GP of the seeds of Chuan 3A and 4A×other B lines were higher than those of Chuan 3A×3B and chuan 4A×4B;although the GW of Chuan 4A and 6A×R were heavier than those of Chuan 4A and 6A×Chuan 7B/R,the GE and GP of the latter crosses increased largely and their RPHScs were less.Therefore,it was considered as effective ways for improvement on seed viability,that A lines cross with other B lines having different genetic background but similar agronomic characters and that the gene(s) concerning high seed viability were transferred into restorers.     

Effects of water stress on male gametophyte development in plants

 Male reproductive development in plants is highly sensitive to water deficit during meiosis in the microspore mother cells. Water deficit during this stage inhibits further development of microspores or pollen grains, causing male sterility. Female fertility, in contrast, is quite immune to stress. The injury is apparently not caused by desiccation of the reproductive tissue, but is an indirect consequence of water deficit in the vegetative organs, such as leaves. The mechanism underlying this stress response probably involves a long-distance signaling molecule, originating in the organs that undergo water loss, and affecting fertility in the reproductive tissue, which conserves its water status. Much research has been focused on the involvement of abscisic acid in this regard, but the most recent evidence tends to reject a role for this hormone in the induction of male sterility. Stress-induced arrest of male gametophyte development is preceded by disturbances in carbohydrate metabolism and distribution within anthers, and an inhibition of the key sugar-cleaving enzyme, acid invertase. Since invertase gene expression can be modulated by sugar concentration, it is possible that decreased sugar delivery to reproductive tissue upon inhibition of photosynthesis by stress is the signal that triggers metabolic lesions leading to failure of male gametophyte development. Received: 31 October 1996 / Revision accepted: 18 February 1997     

Genetics and cytology of a new genic male-sterile soybean [Glycine max (L.) Merr.]

 Genetic and cytological studies were conducted with a new male-sterile, female-fertile soybean [Glycine max (L.) Merr.] mutant. This mutant was completely male sterile and was inherited as a single-recessive gene. No differences in female or male gamete transmission of the recessive allele were observed between reciprocal cross-pollinations in the F₁ or F₂ generations. This mutant was not allelic to any previously identified soybean genic male-sterile mutants: ms1, ms2, ms3, ms4, ms5, or ms6. No linkage was detected between sterility and flower color (W1 locus), or between sterility and pubescence color (T1 locus). Light microscopic and cytological observations of microsporogenesis in fertile and sterile anthers were conducted. The structure of microspore mother cells (MMC) in male-sterile plants was identical to the MMCs in male-fertile plants. Enzyme extraction analyses showed that there was no callase activity in male-sterile anthers, and this suggests that sterility was caused by retention of the callose walls, which normally are degraded around tetrads at the late tetrad stage. The tapetum from male-sterile anthers also showed abnormalities at the tetrad stage and later stages, which were expressed by an unusual formation of vacuoles, and by accumulation of densely staining material. At maturity, anthers from sterile plants were devoid of pollen grains. Received: 13 May 1996 / Revision accepted: 19 August 1996     

Generation of heteroplastidic Nicotiana cybrids by protoplast fusion: analysis for plastid recombinant types

Summary Protoplasts of a mutant line of Nicotiana tabacum having a maternally-transmitted chlorophyll deficiency were fused with protoplasts of two alloplasmic-male-sterile Nicotiana lines by the donor-recipient technique. In both fusion experiments variegated plantlets were regenerated which were shown to contain cytoplasms of mixed chloroplast nature. This confirms that with the donor-recipient method one can obtain mixed cytoplasms of genetically different chloroplasts. We present a convenient system to assay for genetic recombination between chloroplasts by combining use of several cytoplasmic markers: vis. chlorophyll pigmentation, chloroplast DNA restriction patterns, tentoxin resistance and male sterility. Within the limits of the experiment no recombinant types were recovered.     

Lycopersicon esculentum: Trifoliate plants recovered from anther cultures of heterozygous tftf plants

The effects of the genotype and growth medium composition on callus induction and shoot regeneration from tomato (Lycopersicon esculentum Mill) anthers were studied. Five male sterile varieties, homozygous for the recessive gene ms 10³⁵, their isogenic fertile counterparts, and nineteen sterile mutants from an F₂ population segregating for ms 10³⁵, were tested. Callus induction and shoot formation were found to be affected by the genotype. The presence of the mutant gene ms 10³⁵ greatly increased callus induction. A significant interaction concerning callus induction was found between the ms 10³⁵ gene and the general genetic background. In most of the plants shoot regeneration from the anthers was associated with various degrees of callus production. However, there was no correlation between callus production and the ability to regenerate plants from that callus. Anthers isolated from plants which were heterozygous for the recessive leaf marker trifoliate, regenerated diploid plants with trifoliate leaves. The plants retained the trifoliate phenotype for over six months in culture under non-aseptic condition. Since the trifoliate phenotype appears only in the homozygous recessive state, the evidence that these trifoliate plants are doubled haploids of sporogenic origin is discussed.     

Hydroxycinnamic acid amides in fertile and cytoplasmic male sterile lines of maize

Hydroxycinnamic acid (HCA) amides in fertile and cytoplasmic male sterile lines of maize were determined in reproductive organs, developing grains and cobs. HCA amides occurred in large amounts in the anthers of fertile plants (line F7N) and were absent from the anthers of cytoplasmic male sterile lines (lines F7T and F7C). Restoration of fertility was associated with the production of these compounds (line FC31). Considerable variations were observed in the concentrations of HCA amides at different stages of growth and grain maturation. Changes of HCA amides in the grains which were to produce sterile plants followed a pattern similar to that obtained with the grains which were to produce fertile plants. Accumulation of HCA amides was substantially higher in fertile lines whatever their genotype (F7N, FC31 and F7T x FC31) than in sterile lines. Marked changes occurred in the HCA amide content of embryo and endosperm during grain development. Many changes in HCA amides were observed in cobs during development and maturation, but no substantial differences could be observed between fertile and sterile lines.