玛拉巴石斑鱼工厂化人工育苗技术(下)

(2)轮虫、蒙古裸腹蚤的培养:培养池为常用水泥池,24小时连续充气,充气量以轮虫、蒙古裸腹蚤不因缺氧而浮于水面为宜。每天投喂适量酵母和小球藻液,轮虫培养每天投喂酵母量为0.005克/万个,分4～6次投喂;蒙古裸腹蚤投喂酵母量为1～2毫克/升,分3次投喂;每天换水20%,换水后添加小球     

Improvement of Appearance Traits of elite Rice Zhenshan 97B by Molecular Introgression of Targeted about 800 kb Interval from Indica Rice Jiafuzhan Genome

Rice appearance trait is mainly mean grain length (GL), grain width (GW), ratio of length and width (L/W), chalkiness (CH) and transparence (TR). Appearance is the first impression, and rice appearance traits effect significantly the value of commodity. Hybrid rice is relatively low quality, especially on rice appearance quality,which restricts partially its extend and badly export of hybrid seed and rice in China.Shanyou 63, the most famous hybrid rice combination of China, could not be also sustainable application because of its poor qualities. In this studies, We attempt to improve appearance quality of the elite rice maintainer of Zhenshan 97B by using molecular approach of genomic interval targeted introgression. Three F2 populations were constructed to study the classical and molecular genetic of rice appearance traits, at the same time, molecular marker-asisted toimprove the main maintainer line, Zhenshan 97B were performed. There some following results:Three F2 populations were constructed based on crosses of Guangluai 4×Jiafuzhan (GJ popu.), Minghui 86×Jiafuzhan (MJ popu.) and Zhenshan 97B×Jiafuzhan (ZJ popu.). ZJ popu and GJ popu were produced to reconstruct the linkage maps related with rice appearance traits based on BSA, and detect QTL of target traits based on IM.In ZJ popu, 9 QTLs of target traits were detected. The interval, RM169～RM516 on Chromosome 5, was genetic locus of GW, L/W, CH and BW, and the explained variances were 10.9%, 14.9%, 12.0% and 14.2%, respectively.RM214 on chromosome 7, RM339 on chromosome 8 were genetic loci if CH and BW, and the explained variances were 9.4%, 10.0%, 11.0% and 12.1%. besides there is a genetic locus, RM347, of G/W on chromosome 3, and the explained varance was 10.3%. In GJ popu, 5 QTLs of rice appearance traits were detected. The interval, RM169-RM516 on Chromosome 5, were also the genetic loci of GW, CH and BW, and the explained variances were 9.0%, 12.5%, and 13.6%, respectively. RM264～RM80 on chromosome 8 was the genetic loci of GW amd L/W,and the explained variances were 14.7% and 13.4%. The genotypes of all QTLs, except RM214, from Jiafuzhan contributed to improving riee appearance traits.The research was to improve the rice appearance quality of Zhenshan 97B by introgressing QTLs of rice appearance traits from Jiafuzhan, a donor parent, by means of MAS in the process of recurrent backcrossing, 4 SSR markers were used to detect the target genotypes from donor parent, and 70 SSR primers to identify the genetic background of recurrent parent. Consequently, there were 19 target individuals in BC3, in which the mean value of GL, GW, L/W, CH and BW were 6.06±0.04mm, 2.50±0.02mm, 2.43±0.02, 29.5±2.7 and 29.4±2.7%, respectively,and the mean homologous degree (HD) to recurrent parent is up to 98.22%. Compared to Zhenshan 97B, GL, CH and BW decreased 0.17 mm, 34.9% and 34.5% respectively and however L/W increased 0.19. which accounted separately for 25.37%, 54.79%, 54.57% and 10.50% of thedifference of every trait between two parents. It is very luckly to find a individual, 13BC2-3BC3 line, whose GL, GW, G/W, CH and BW were 6.04 mm, 2.43 mm, 2.49,2.5% and 2.5% separately, There is very important signification for it to be used in hybrid rice breeding improving appearance traits. In the research, MAS efficiency of 3 loci (with 4 markers ) was 5.68% and coincident degree was 73.08%.     

Improving Drought Tolerance of Rice by Designed QTL Pyramiding

Drought is the most important factor limiting rice yields in the rainfed areas of Asia. To overcome the problem, we developed a new strategy ＇designed QTL pyramiding＇ to more efficiently develop drought tolerant （DT） rice cultivars. In the first phase of this strategy, we developed large numbers of introgression lines （ILs） in elite backgrounds using backcross （BC） breeding, each of which carries multiple genomic segments for improved DT from a known donor. Then, we genotyped all ILs with SSR markers to track the genomewide pattern of introgression in the DT ILs of 16 BC populations derived from crosses between 2 recipients （IR64 and Teqing） and four different donors. X^2 tests and linkage disequilibrium analyses of introgression in the DT ILs revealed significant frequency shifts at many loci across the genome and non-random associations at the multilocus level as a result of selection for DT in the ILs, which led us to the discovery of putative genetic networks underlying DT in the ILs. The networks each containing all DT loci detected in ILs from a BC population showed some interesting features.     

Cloning and Sequence Analysis of Lipoxygenase Gene cDNA from Cucumber Fruit （Cucumis sativus L.）

Lipoxygenases are nonheme-iron-containing dioxygenases that catalyze the hydroperoxidation of unsatrated fatty acids containing a cis, cis-1,4-pentadiene structure producing hydroperoxy acids with conjugated dienes. LOX activity has been found in a wide range of plants. Typical substrates for LOX in plants are linoleic acid and linilenic acid fatty acids. The function of various LOXs in plants is unknown, but their participation in all stages of plant growth and development has been suggested （Hildebrand, 1989; Siedow, 1991）. Some of the physiological processes in whicn lipoxygenses have been implicated include wounding （Saravitz and Siedow, 1996）, pathogen attack （Melan et al., 1993）, seed germination （Kato et al., 1992）, fruit ripening （Ferrie et al., 1994）, plant senescence （Paliyath and Droillard, 1992）. The study on the role of lipoxygenase in ripening and senescence fruit focused on tomato and strawberry. Cloning LOX gene of cucumber fruit will make us further understand the molecularaction mode of this enzyme during fxuit ripening and senescence. In this paper we isolated the partial nucleotide sequences of cucumber fiuit lipoxygenase gene and discuss the characterization of it.     

Gene Discovery and Characterization from Non-sequenced Medicinal Plants

Plant-derived secondary metabolites still constitute important resources for currently prescribed drugs and for the discovery of active principles which are effective in new indication areas. Promising compounds are often exclusively found in wild medicinal plants and the classical plant-based pharmaceutical production approach has been hampered by low yields, lack of reproducibility and the high complexity of plant extracts （Rischer and Oksman-Caldentey, 2005）. Influencing the accumulation of target compounds by breeding or genetic engineering is greatly impeded because in most cases the general biosynthetic routes are incompletely understood.     

TILLING and Ecotilling the Plant Kingdom

The modem crop scientist has a large amount of available nucleotide sequence information to identify genes of potential agronomic importance. Using reverse genetic approaches, specific genes can be disrupted, and hypotheses regarding gene function directly tested in vivo. Although a number of reverse genetic methods have been introduced, many are limited in application because they are organism-specific, expensive, transgenic, or only transiently disrupt gene function. However, traditional mutagenesis using chemical mutagens has been widely used as a forward genetics strategy to create many new crop plant varieties at relatively low cost. Mutagens such as ethyl methanesulphonate （EMS） cause stable point mutations and thus produce an allelic series of truncation and missense changes that can provide a range of phenotypes （Greene et al., 2003）. TILLING （targeting induced local lesions IN genomes） is a high-throughput reverse genetic strategy that combines traditional mutagenesis and SNP discovery methods （Colbert et al., 2001; McCallum et al., 2000）. To identify mutations, target regions of-l.5 kb are amplified with fluorescently labeled gene specific primers. Heteroduplexes are then formed between wild-type and mutant strands, mismatches are cleaved by incubation with a single-strand specific nuclease,     

Mapping QTLs for Yield Stability in Durum Wheat Grown under Different Water Regimes

Among the most important Mediterranean annual crops, durum wheat is widely grown in drought-prone areas. Therefore, improving water-use efficiency （WUE） of durum wheat represents a major breeding goal. IDu-WUE （Improving Durum wheat for Water Use Efficiency and yield stability through physiological and molecular approaches） is a collaborative project among public and private research centres in Italy, Spain and WANA （West Asia and North Africa） countries （Morocco, Tunisia, Syria and Lebanon） funded by the European Union aimed at investigating the genetic variation for WUE and yield stability in durum wheat grown in Mediterranean droughtprone areas. During the first year of the project, a number of morpho-physiological traits （e.g. early vigour, flowering time, leaf rolling, number of fertile tillers, etc.）, WUE, WUE-related traits （e.g. carbon isotope discrimination, canopy temperature, chlorophyll fluorescence, etc.）, yield and its components have been investigated in a RIL population （249 lines） and a collection of ca. 190 durum wheat accessions characterized by a high level of linkage disequilibrium （Maccaferri et al., 2005）,     

A Simple and Reliable Assay for Detecting Specific Nucleotide Sequences in Plants Using Optical Thin-film Biosensor Chips

Here we report the adaptation and optimization of an efficient, accurate and inexpensive assay that employs custom-designed silicon-based optical thin-film biosensor chips to detect unique transgenes in genetically modified （GM） crops and SN-P markers in model plant genomes. Briefly, aldehyde-attached sequence-specific singlestranded oligonucleotide probes are arrayed and covalently attached to a hydrazine-derivatized biosensor chip surface. Unique DNA sequences （or genes） are detected by hybridizing biotinylated PCR amplicons of the DNA sequences to probes on the chip surface. In the SN-P assay, target sequences （PCR amplicons） are hybridized in the presence of a mixture of biotinylated detector probes and a thermostable DNA ligase. Only perfect matches between the probe and target sequences, but not those with even a single nucleotide mismatch, can be covalently fixed on the chip surface. In both cases, the presence of specific target sequences is siL, nified by a color change on the chip surface （gold to blue/purple） after brief incubation with an anti-biotin IgG horseradish peroxidase （HRP） to generate a precipitable product from an HRP substrate.