海参i型溶菌酶基因及其编码产物的结构特点

通过RT-PCR 和 RACE PCR技术,从海参(Stichopus japonicus)体壁中克隆得到一种溶菌酶基因(GenBank：EF036468).生物信息软件分析表明,其中全长cDNA为 713 bp,5′非编码区（UTR）246 bp,3′UTR 29 bp,开放阅读框438 bp,编码145个氨基酸,包括溶菌酶成熟肽124个氨基酸和信号肽21个氨基酸.对海参溶菌酶与多种无脊椎动物的c、g和i型溶菌酶进行分析比较,发现它与i型溶菌酶有较高的同源性,并具有i型溶菌酶高度保守的2个活性位点,即Glu34和Ser50.活性位点附近具有i型溶菌酶的一段特有的氨基酸保守序列MDVGSLSCG(P/Y)(Y/F)QIK,所以推断克隆的海参溶菌酶为i型.另外,通过搜索蛋白保守结构域数据库,发现海参溶菌酶与医用水蛭失稳酶相似性最高,并且这2个酶的三级结构模型也极其相似.因此推测,海参i型溶菌酶具有双功能特性,既能作用于细菌细胞壁的糖苷键使细胞裂解,又具有失稳酶的一些生化功能,能够水解纤维蛋白,这些特点在海参自溶过程中发挥重要的作用.     

两种海马鳃组织和消化道的黏液细胞类型及分布

运用阿新兰(AB,pH 2.6)和过碘酸雪夫氏(PAS)反应染色方法,对三斑海马(Hippocampus trimaculatus)和日本海马(H.japonicus)鳃组织与消化道中的黏液细胞类型及分布进行了研究。染色结果显示:两种海马的鳃组织和消化道中均含有黏液细胞,日本海马的鳃组织中含有Ⅰ型和Ⅳ型黏液细胞,三斑海马的鳃组织中含有Ⅰ型、Ⅲ型和Ⅳ型黏液细胞。两种海马消化道各部位的黏液细胞类型和数量有明显差异:日本海马的食道中Ⅰ型细胞最多,而三斑海马的食道中Ⅳ型细胞最多;日本海马的前肠中只含有Ⅰ型细胞,而三斑海马的前肠中含有Ⅰ型、Ⅲ型和Ⅳ型细胞,其中Ⅰ型细胞含量最多;日本海马的中肠中含有Ⅰ型、Ⅲ型和Ⅳ型细胞,其中Ⅲ型细胞含量最多,而三斑海马中肠中只含有Ⅰ型细胞;日本海马与三斑海马的后肠中都分布有Ⅰ型、Ⅱ型、Ⅲ型和Ⅳ型细胞,两者不同的是,日本海马的后肠中Ⅲ型细胞含量最多,三斑海马的后肠中Ⅳ型细胞含量最多。     

Classification,inhibition, and specificity studies of the vitelline coat lysin from toad sperm

The sonicated supernatant of the sperm of the toad, Bufo japonicus, can digest easily the vitelline coat (VC) of uterine eggs, and to a lesser extent the VC of coelomic eggs, but not that of activated eggs. The VC lysis and fertilization were competitively inhibited in the presence of t-butyloxycarbonyl-L-Gln-L-Arg-L-Arg-4-methylcoumaryl-7-amide (Boc-Gln-Arg-Arg-MCA), suggesting the involvement of proteases in the fertilization process. Starting from a sonicated supernatant, a potent VC lysin, possessing hydrolytic activity on Boc-Gln-Arg-Arg-MCA, was obtained by anion-exchange chromatography and gel filtration. The activity of the partially purified lysin was inhibited by diisopropyl fluorophosphate (DFP) and by such trypsin inhibitors as soybean trypsin inhibitor, leupeptin, and (p-amidinophenyl) methanesulfonyl fluoride hydrochloride, but not by chymostatin, E-64, and ethylene glycol bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid. The molecular weight of the lysin was estimated to be 32K, based on the fluorographic image of ³H-DFP binding to the lysin on sodium dodecyl sulfate gel electrophoresis. The VC lysin was most active at pH 7.0–7.6 and under low ionic strength equivalent to fresh water. The release of the VC lysin was induced upon incubation of sperm with the contents of oviducal pars recta granules (PRG), which are known to induce the acrosome reaction. We conclude that the protease studied here represents the VC lysin of toad sperm that is involved in fertilization by digesting the VC of uterine eggs, probably released as a result of the acrosome reaction induced by PRG.     

Leucocyanidin reductase activity and accumulation of proanthocyanidins in developing legume tissues

Proanthocyanidin (PA) and anthocyanin accumulation and location in developing leaves, flowers, and seeds of the legumes Medicago sativa, Lotus japonicus, Lotus uliginosus, Hedysarum sulfurescens, and Robinia pseudacacia were investigated by quantitative measurements and by histological analysis after staining with 1% vanillin/HCl, butanol/HCl, or 50% HCl. M. sativa leaves and flowers, L. japonicus leaves, and R. pseudacacia flowers do not contain PAs, but seeds of all investigated species contain PAs. Anthocyanins are absent in the seed coats of all five species and in leaves of L. japonicus. PA content generally increases as a function of development in leaves, but declines in flowers. With the exception of H. sulfurescens, flower PAs are synthesized in the parenchyma cells of the standard petal, while anthocyanins are located in the neighboring epidermal cells. Leucocyanidin reductase (LCR) catalyzes the conversion of 2,3-trans-3,4-cis-leucocyanidin to (+)-catechin and is the first enzyme in the PA-specific pathway. LCR activity was only detected in PA-containing tissues and generally declined during tissue development.     

Hatch date‐dependent differences in early growth and development recorded in the otolith microstructure of Trachurus japonicus

Otolith microstructure of 308 juvenile jack mackerel Trachurus japonicus sampled in the southern Sea of Japan between June and September 2002 were examined by light microscopy. Hatch‐date distribution indicated a protracted spawning season from 16 January to 30 May 2002. The first secondary primordium (SP) was formed at ages 19–54 days, with a mean ±  s . d . of 30·4 ± 6·1. The number of SP in an otolith ranged from two to 15, with a mean of 6·1 ± 1·7. Age of the first SP formation, number of SP and increment width varied with hatch date. In general, late‐hatched fish were younger when the first SP formed, and had a higher number of SP and wider increments, than early‐hatched fish. As increment width is a linear function of somatic growth and formation of the first SP is associated with metamorphosis in jack mackerel, these variations in otolith microstructure indicated that rates of growth (as inferred from wider increments) and development (as inferred by younger age of first SP formation) were higher in late‐hatched fish relative to early‐hatched fish. Higher temperature experienced by late‐hatched fish during early life stages may contribute critically to their higher growth and development rates compared to early‐hatched fish.     

百脉根BIO和豌豆突变位点ELE2的比较基因组定位

豆科两侧对称花的花瓣具有背腹(DV)的分化以及可变的器官内部(IN)非对称性,在大小与形状上显示出不同的发育特征;因而花瓣的发育为克隆决定植物器官的形状与大小的关键基因提供了很好的实验系统。本研究对百脉根中BIO基因进行研究。百脉根bio突变体具有多效性,既影响花器官内部的对称性也影响器官的大小和育性,豌豆ele突变体的表型与bio相似。定位结果表明BIO和ELE2位于豆科基因组的共线性区段,提示BIO和ELE2可能是同源基因突变所致。本研究利用比较基因组定位方法,将BIO和ELE2候选基因锚定在豆科模式植物百脉根和蒺藜苜蓿基因组含有11个同源基因的BAC重叠群上。BIO和ELE2基因的克隆将有助于揭示豆科花瓣形态和大小调控的分子机理,进而为豆科作物遗传改良提供分子理论基础。     

百脉根嫁接方法的优化及其在结瘤自调控研究中的应用

嫁接试验是研究结瘤自调控的有效方法。本文优化了百脉根的嫁接方法,并应用该方法初步研究了百脉根突变体rel3根瘤数目减少的机理。结果表明,rel3突变体根瘤数目减少的表型是由来自于茎的信号决定。     

铀尾矿库区的植物组成与多样性

对华中南某铀尾矿库区的植物组成及多样性进行了研究,并测定了各样地环境辐射状况。结果显示,库区有高等植物79种,隶属32科67属。不同环境辐射强度生境上植物重要值(IV)差别较大:轻度(LR)、中度(MR)辐射强度生境的群落物种重要值相对分散,乔灌草3层均出现多个优势植物;而重度(HR)辐射强度群落的优势植物集中于极少数种上。结合物种重要值、群落相似性系数(C)及聚类分析可把库区植物群落分为4类:1)轻度辐射(LR)的樟树+构树—红继木+牡荆—博落回+商陆群落;2)中度辐射(MR)的苦楝+构树—牡荆+白背桐—博落回+五节芒群落;3)白背桐—芦苇+白茅灌丛和4)盐肤木—狗尾草+白茅+圆果雀稗灌丛,分布于重度辐射(HR)。各样地的Shannon多样性指数(H)为0.463～1.734,表现为轻度中度重度;Margalef丰富度(DMA)为33.962～49.024,Pielou均匀度指数(Jsw)为0.463～0.986。文中还分析库区存在的生态环境问题。     

A genetic linkage map of the sea cucumber, Apostichopus japonicus (Selenka), based on AFLP and microsatellite markers

We present the first genetic maps of the sea cucumber ( Apostichopus japonicus ), constructed with an F₁ pseudo-testcross strategy. The 37 amplified fragment length polymorphism (AFLP) primer combinations chosen identified 484 polymorphic markers. Of the 21 microsatellite primer pairs tested, 16 identified heterozygous loci in one or other parent, and six were fully informative, as they segregated in both parents. The female map comprised 163 loci, spread over 20 linkage groups (which equals the haploid chromosome number), and spanned 1522.0 cM, with a mean marker density of 9.3 cM. The equivalent figures for the male map were 162 loci, 21 linkage groups, 1276.9 and 7.9 cM. About 2.5% of the AFLP markers displayed segregation distortion and were not used for map construction. The estimated coverage of the genome was 84.8% for the female map and 83.4% for the male map. The maps generated will serve as a basis for the construction of a high-resolution genetic map and mapping of the functional genes and quantitative trait loci, which will then open the way for the application of a marker-assisted selection breeding strategy in this species.     

麦冬花药绒毡层和乌氏体的细微结构

麦冬(Ophiopogon japonicus)的绒毡层发育为分泌型。在小孢子母细胞时期,绒毡层细胞达到了发育的高峰。此时,绒毡层细胞中细胞器非常丰富,具大量线粒体、高尔基体和质体,尤以肉质网含量最多;原乌氏体出现较早,在小孢子母细胞时期绒毡层细胞中就已出现;四分体时期,大量原乌氏体被排入内切向面的质膜和纤维素壁之间;到了小孢子早期,绒毡层细胞失去细胞壁,原乌氏体分布在质膜的凹陷处,孢粉素物质在其上沉积,发育为乌氏体,乌氏体有单个和复合两种类型;当花粉成熟时,绒毡层细胞完全解体。