主要禾谷类作物比较基因组学研究策略与进展

随着拟南芥、水稻等模式植物基因组测序计划的完成, 比较基因组学作为一门新兴学科, 近年来发展迅速, 为植物基因组的进化、结构和功能研究开辟了新的途径。文章综述了比较基因组学在作物比较遗传作图、基因结构区域的微共线性、ESTs和蛋白质水平的比较以及基于比较基因组学的基因和QTL的克隆等方面内容与研究进展, 分析了不同水平上比较基因组学研究策略的原理、特点、可行性, 以期为利用模式生物的基因和基因组数据、采用比较基因组学策略克隆作物重要性状功能基因、阐明基因组结构与进化提供帮助。     

植物转基因整合座位的结构

通过农杆菌和直接DNA转移技术所获得的转基因植株都具有复杂的转基因座位, 转基因整合染色体和染色体区段是随机的, 但组织培养的选择作用表现为非随机性, 偏向整合于染色体的基因富集区。转基因座位除少数含有完整的单拷贝转基因之外, 大多数转基因座位中外源转基因片段、基因组片段和填充DNA相间而存在。转基因座位中转基因及基因组DNA片段产生缺失、重复和染色体的重排, 转基因的完整性对转基因表达具有重要作用。     

Genetic Association Between Ca2+-ATPase Activity and Audiogenic Seizures in Mice

Ca2+-ATPase activity was studied in fresh brain stem homogenates of the audiogenic seizure (AGS)-resistant C57BL/6 and AGS-susceptible DBA/2 inbred strains and in 21 B6 X D2 recombinant inbred strains. A highly significant negative correlation was found between Ca2+-ATPase activity and AGS susceptibility among these strains. In general, strains with low Ca2+-ATPase activities were more AGS-susceptible than strains with high activities. Further, Ca2+-ATPase activity appears to be influenced by a major gene associated with the Ah locus. This gene is designated Caa for Ca2+-ATPase activity and is different from Ias, which is closely linked to the Ah locus. Ias influences AGS spread by a yet unknown biochemical mechanism, whereas Caa may influence AGS susceptibility by regulating Ca2+-ATPase activity in brain tissue.     

The base-alteration spectrum of spontaneous and ultraviolet radiation-induced forward mutations in the URA3 locus of Saccharomyces cerevisiae

Summary A forward mutation system has been developed to obtain rapidly clonable mutants at the URA3 locus in yeast by means of selection for 5-fluoroorotic acid resistance. We have used this system to determine base changes in 35 spontaneous and 34 ultraviolet radiation-induced ura3 base substitution mutants. Other mutants (frameshift, deletion, duplication, replacement) were detected as well. Evidence is reported which suggests cyclobutane dimers are the principal mutagenic lesions induced by UV radiation in stationary phase cells of the yeast Saccharomyces cerevisiae. Since most of the induced lesions are at 5-TT-3 sites, the results suggest that the A-rule, preferential insertion of adenine residues opposite poorly pairing sites in DNA, does not apply for yeast cells irradiated in stationary phase, whereas the spontaneous mutation data indicate that the A-rule applies for cells in logarithmic phase. Most of the spontaneous mutations are transversions. UV-induced transitions and transversions occur at approximately equal frequencies.     

Insertional DNA and spontaneous mutation at the white locus in Drosophila simulans

A large body of data on molecular analyses of several multiallelic loci in Drosophila melanogaster has demonstrated a high incidence of mobile DNA element insertions among spontaneous mutations. In the sibling species D. simulans, the dispersed, middle repetitive, nomadic sequences are reduced to about one-seventh that of its sibling species (Dowsett and Young 1982). Does this reduced amount of middle repetitive DNA (or mobile DNA sequences) mean that in D. simulans the occurrence of insertion mutants will be rare compared with that of D. melanogaster? To test this possibility, we collected seven different spontaneous white mutants of D. simulans and studied their molecular gene structures. Five out of seven mutants had insertion sequences which varied in length from 0.4 kb to 16 kb. One bore a deletion spanning the w region and another showed no gross structural alteration. Thus the proportion of insertional mutations at the white locus in D. simulans is equivalent to that observed in D. melanogaster. Among the five insertional mutants, one, wmky, showed genetic instability; the other four were stable. wmky was found to mutate at a frequency of 2.1 x 10(-5) in meiotic cells and may also be unstable in somatic cells.     

cDNA,amino acid and carbohydrate sequence of barley seed-specific peroxidase BP 1

The major peroxidase of barley seed BP 1 was characterized. Previous studies showed a low carbohydrate content, low specific activity and tissue-specific expression, and suggested that this basic peroxidase could be particularly useful in the elucidation of the structure-function relationship and in the study of the biological roles of plant peroxidases (S.K. Rasmussen, K.G. Welinder and J. Hejgaard (1991) Plant Mol Biol 16: 317–327). A cDNA library was prepared from mRNA isolated from seeds 15 days after flowering. Full-length clones were obtained and showed 3 end length variants, a G+C content of 69% in the translated region, a 90% G or C preference in the wobble position of the codons and a typical signal peptide sequence. N-terminal amino acid sequencing and sequence analysis of tryptic peptides verified 98% of the sequence of the mature BP 1 which contains 309 amino acid residues. BP 1 is the first characterized plant peroxidase which is not blocked by pyroglutamate. BP 1 polymorphism was observed. BP 1 is less than 50% identical to other plant peroxidases which, taken together with its developmentally dependent expression in the endosperm 15–20 days after flowering, suggests a unique biological role of this enzyme. The barley peroxidase is processed at the C-terminus and might be targeted to the vacuole. The single site of glycosylation is located near the C-terminus in the N-glycosylation sequon -Asn-Cys-Ser- in which Cys forms part of a disulphide bridge. The major glycan is a typical plant modified-type structure, Man1-6(Xyl1-2)Man1-4GlcNAc1-4(Fuc1-3)GlcNAc. The BP 1 gene was RFLP-mapped on barley chromosome 3, and we propose Prx5 as the name for this new peroxidase locus.     

谷氨酸钠兴奋室旁核引起的升压效应及其与蓝斑-升压反应和A_1-降压反应间的关系

将L-谷氨酸钠(Glu)注入乌拉坦麻醉、箭毒化、人工呼吸大鼠的室旁核(NPV)或蓝斑内引起升压反应。蓝斑升压反应可被双侧室旁核内预先注射酚妥拉明或心得安明显衰减;双侧室旁核内预先注射酚妥拉明或荷包牡丹碱还可使Glu兴奋延髓A_1区引起的降压反应减小,但注射心得安对A_1-降压反应无明显影响。以上结果提示蓝斑-升压反应和A_1-降压反应均部分通过NPV实现,A_1降压过程中可能有NPV内GABA能抑制性中间神经元参与。     

中枢去甲肾上腺素能系统对大鼠颈动脉窦反射的影响

孤离颈动脉窦,向侧脑室(LCV)注射6-羟多巴胺(6-OHDA,200μg)和电解损毁蓝斑(LC),建立窦内压(ISP)-平均动脉压(MAP)关系曲线,与对照组比较,研究中枢去甲肾上腺素能系统对ISP调节MAP的影响。通过Logistic曲线方程符合所有ISP-MAP曲线,曲线的特征是由两个参数即曲线斜率的反演点坐标和MAP变动范围决定的,MAP变动范围同斜率因子的变化率有关,并受ISP的控制。结果如下:(1)注射6-OHDA后,ISP-MAP曲线斜率变化率显著低于对照组,窦内压控制的平均动脉压变动范围明显缩小,而曲线斜率反演点的横坐标(即ISP的位置)无改变。(2)与对照组相比,电解损毁LC后,引起ISP-MAP曲线相似于注射6-OHDA后的变化,但不如后者显著。结果提示:中枢去甲肾上腺素能神经系统可易化颈动脉窦反射。     

Reduction to homozygosity is the predominant spontaneous mutational event in cultured human lymphoblastoid cells

Expression of recessive mutant phenotypes can occur by a number of different mechanisms. Inactivation of the wild-type allele by base-substitution mutations, frameshift mutations or small deletions occurs at both hemizygous and heterozygous cellular loci, while other events, such as chromosome level rearrangements, may not be detected at hemizygous loci because of inviabiltty of the resulting mutants. In order to assess the relative contribution of each type of mutational event, we isolated a human lymphoblastoid cell line that is heterozygous at the adenine phosphoribosyltransgerase (aprt) locus. The mutation rate for the expression of the mutant phenotype (aprt^+/−→aprt^−/−) was 1.3 × 10⁻⁵/cell/ generation. Molecular analysis of the DNA from 26 mutant clones revealed that 19% had undergone deletion of the entire wild-type allele. The aprt heterozygote carries a mutation in the coding sequence of the gene that results in the loss of a restriction site. Analysis of aprt^−/− mutants for this restriction fragment length difference reveales that 23% of the mutants contained point mutations or small ((< 100 bp) deletions. The remainder of the mutants (58%) resulted from reduction to homozygosity of the mutant allele. We suggest that, as in tumor cells in vivo, reduction to homozygosity is a major mechanism for the expression of recessive mutant phenotypes in cultured human cells.