Use of isozyme patterns in characterization of potato mutant lines selected for agronomic quantitative traits

Summary Seven varieties and 57 spontaneous or induced in vitro mutant lines (20 macromutant and 37 micromutant events) of potato were tested by starch gel electrophoresis for ADH, GOT, PGI, PGM, ACO, IDH, MDH and 6PGDH isozymes in tuber extracts. The data showed that in contrast to variety comparisons, the isozyme patterns rarely differentiate mutant lines which have altered morphological traits. But trying to identify isozyme differences in mutants can still be useful for a chimeric structure for GOT-2 alleles in a mutant from Atlantic and a new tuber specific locus for 6PGDH in mutants from Russet Burbank were found.Abbreviations ACO aconitase - ADH alcohol dehydrogenase - GOT glutamate oxaloacetate transaminase - IDH isocitric acid dehydrogenase - MDH malate dehydrogenase - 6PGDH 6-phosphogluconate dehydrogenase - PGI phosphoglucoisomerase - PGM phosphoglucomutase - SGE starch gel electrophoresis - EMS ethyl metanesulfonate     

Resistance to Meloidogyne incognita in Ethiopian Coffea arabica accessions

Polyacrylamide gel electrophoresis (PAGE) was used to study the polymorphism of esterases in Allium cepa L. and A. fistulosum L. Two varieties of each species, an A. fistulosum × A. cepa interspecific F₁ hybrid, and (A. fistulosum × A. cepa) hybrid derivatives were analyzed for determination of banding patterns upon staining with α- and β-napthyl acetate substrates of esterase. Complex band patterns were observed. In total, 10 bands were detected between A. cepa and A. fistulosum — five inA. cepa, six in A. fistulosum with only one band common to both species. With the exception of one band unique to A. fistulosum which appeared only when stained with α-substrate, extracts of both A. cepa and A. fistulosum leaf tissue exhibited the same bands when stained with both α- and β-substrates. Bands stained with the different systems are distinguished by color: α-substrate always appeared black, while bands stained to β-substrate are always red. Esterase bands were assigned into 5 presumptive loci of four zones of activity with according to the migration distance of the bands from the front, color of each band upon staining with α- and β-substrates, and segregation observed in crosses and hybrid derivatives. Esterase enzymes detected in this study appear to be monomeric. Polymorphisms were identified between A. cepa and A. fistulosum by esterase banding patterns. Esterase enzymes provide an additional marker in monitoring introgression of foreign germplasm in interspecific onion breeding. This revised version was published online in August 2006 with corrections to the Cover Date.     

An improved acetic acid-urea polyacrylamide electrophoresis method to evaluate bovine sperm protamines

The acetic acid-urea polyacrylamide gel electrophoresis system could separate very similar basic proteins on differences in size and effective charge. This system has been used for many years to analyse histones and their post-translational modifications and widely used in the study of mammal protamines. Two types of protamine have been described, the protamine 1 (P1) and the protamine 2 (P2) family members, which are synthetized by PRM1 and PRM2 genes. The ratio of P1 and P2 is important for predicting fertility in humans and mice. Therefore, the quantification of protamines is a fundamental step in order to establish the ratio between P1 and P2 in these species. In other mammals, studies linking sperm protamination and the protamine ratio with fertility are increasing. So, the use of an effective technique to separate and quantify protamines is important to study sperm P1/P2 ratio. Therefore, this article describes in detail a feasible and useful procedure to isolate bovine sperm protamines, to perform pre-electrophoresis with PEG solution and finally to carry out acid-urea polyacrylamide gel electrophoresis in reverse polarity. This technique allows a clear separation and efficient detection of bovine sperm protamines.     

改进的ISTA麦醇溶蛋白聚丙烯酰胺凝胶电泳方法在小麦品种真实性和纯度鉴定中的应用

本研究以小麦品种为材料，在ISTA麦醇溶蛋白的标准电泳程序基础上，对其进行了改进，凝胶浓度由10％增加为16．5％．既提高了分辨率，又使得剥肢容易。同时，凝胶交联度由4％降为3．3％，硫酸亚铁的浓度也有所降低，增加了胶的韧性。凝肢溶液中过氧化氢的量经过多次试验，也进行了定量。由原来的少量定量为70出，从而形成了一套适合于一般实验室就可进行小麦品种真实性和纯度鉴定的方法。并将其应用于小麦种子的真实性和纯度的快速鉴定中。     

Characterization of developing microbial communities in Mount St. Helens pyroclastic substrate

The 1980 eruption of Mount St. Helens created a unique opportunity to study microbial communities in a developing soil ecosystem containing little total carbon (C) or total nitrogen (N). We collected surface samples (0-5 cm) from areas near Mount St. Helens National Volcanic Monument 17 years after the eruption. The samples were from bare soil with no plant development, soil under living prairie lupine (Lupinus lepidus) and dead prairie lupine in the pyroclastic plain near Spirit Lake, Washington. We also collected soil from a nearby forested area. Phospholipid fatty acids (PLFAs) from pyroclastic materials were analyzed to determine changes in soil microbial composition. Total bacterial DNA was also extracted from the soils and denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes and DNA sequence analysis of cloned 16S rRNA gene libraries were used to determine the influence of plants on microbial development. Both principal components analysis (PCA) of PLFA fingerprints and non-metric multidimensional scaling (NMS) of DGGE fingerprints distinguished the four soils. Lupine plants influenced the PLFA and DGGE fingerprints depending on the distance of the samples from the plant. DGGE and PLFA profiles from the forest soil were significantly different (P=0.001, based on Monte Carlo permutation test) from those of the bare soil and soil with live lupine. Bacterial clone libraries were constructed, and 800 clones were analyzed by amplified ribosomal DNA restriction analysis (ARDRA) and grouped into operational taxonomic units (OTUs). A total of 51, 77, 58, and 42 different OTUs were obtained from forest soil, soil with live and dead lupine, and bare soil, respectively. Phylogenetic analysis revealed that 62% of the 228 OTUs were classified as Proteobacteria, Actinobacteria, Acidobacterium, Verrucomicrobia, Bacteroides, Cyanobacteria, Planctomycetes, and candidate divisions TM7 and OP10. Members of Proteobacteria represented 29% of the OTUs. Thirty-eight percent of the OTUs could not be classified into known bacterial divisions. This study emphasized the role of prairie lupine in the establishment of pioneering microbial communities and the subsequent roles the biotic components played in improving the quality of pyroclastic soil.     

荞麦、玉米、马铃薯淀粉凝胶特性的试验研究(英)

利用电子万能材料试验机对荞麦、玉米、马铃薯淀粉的力学特性进行了研究。结果表明：在一定范围内，随着淀粉乳浓度的增加，荞麦、玉米、马铃薯的凝胶强度、弹性模量和凝胶弹性呈线性增加，但凝胶弹性变化较小；同一淀粉乳浓度下凝胶强度由高到低顺序为马铃薯淀粉＞玉米淀粉＞荞麦淀粉，弹性模量为马铃薯淀粉＞玉米淀粉＞荞麦淀粉，凝胶弹性为荞麦淀粉＞玉米淀粉＞马铃薯淀粉。在淀粉乳浓度为20％时，随着NaCl浓度增加，3种淀粉的凝胶强度均有一定程度增强。在同一NaCl浓度下，其凝胶强度为马铃薯淀粉＞玉米淀粉＞荞麦淀粉，弹性模量为马铃薯淀粉＞玉米淀粉＞荞麦淀粉，对凝胶弹性的影响不大。     

Overland flow systems for treatment of landfill leachates—Potential nitrification and structure of the ammonia-oxidising bacterial community during a growing season

Overland flow systems are useful for treating landfill leachates, because they provide favourable conditions for nitrification and they are easy to maintain. However, little is known about the microbial communities in such systems or the nitrification capacity of those microorganisms. In this study, seasonal variations in potential nitrification and in community composition of nitrifying bacteria were investigated in two overland flow areas receiving leachate from landfills at Korslöt and Hagby, Sweden. Samples were collected in the settling ponds sediment and at two depths in the overland flow areas (the macrophyte litter layer and the rhizosphere) in May, August and November 2003. A short-term incubation method was used to measure potential oxidation of ammonia and nitrite (designated PAO and PNO). The ammonia-oxidising bacterial (AOB) community was investigated using a 16S rRNA gene approach that included PCR amplification and analysis of PCR products by denaturing gradient gel electrophoresis (DGGE), followed by nucleotide sequencing and phylogenetic analysis.PAO was determined in the range 5-2700 (NO₂⁻+NO₃⁻)-N g⁻¹ dw d⁻¹ and PNO in the range 60-2000 μg NO₂⁻-N g⁻¹ dw d⁻¹. At Korslöt, PAO and PNO showed similar temporal variation in the different ecosystems, whereas no such relationship was noticed at Hagby. Considering both sites, there was no obvious change in the composition of the AOB community over the growing season. However, the composition did differ between the ecosystems: Nitrosomonas-like sequences were more common in the ponds, and in the litter layers they were found as often as Nitrosospira-like sequences, whereas Nitrosospira-like sequences were more common in the rhizospheres. Altogether, we found nine different AOB sequences, five Nitrosomonas-like and four Nitrosospira-like, which belonged to clusters 0, 2, 3b, 6a, 6b and 7. There was no apparent relationship between the number of AOB populations and the PAO in different soil layers and sediments.     

Coupled nitrification-denitrification associated with liquid manure in a gel-stabilized model system

Summary Following the application of liquid manure to soil, the development of the two processes, nitrification and denitrification, was studied in a two-phase model system. A saturated mixture of manure and soil, stabilized with silica gel, was overlain by an aerobic soil phase. Profiles of the redox potential pH, inorganic N, dissolved organic C, nitrification and denitrification potentials, and phospholipid concentrations for an estimate of microbial biomass were measured during a 20-day period. NH ₄ ⁺ diffusing into the aerobic soil was oxidized within 10 mm of the interface, but with only a small accumulation of NO ₂ ^- and NO ₃ ^- . It was estimated that N equivalent to approximately 70% of the NH ₄ ⁺ originally present in the manure was lost through coupled nitrification-denitrification. The potentials for nitrification and denitrification increased 40-and 20-fold, respectively, around the interface. Maximum values were recorded after 14 days. Within 0–5 mm of the anaerobic zone, apparent generation times for NH ₄ ⁺ -oxidizing bacteria of 1.1–1.8 days were estimated between day 1 and day 7. The phospholipid concentration profiles suggested that the biomass within 2 mm on either side of the interface was stimulated throughout the 20-day period.     

DNA- versus RNA-based denaturing gradient gel electrophoresis profiles of a bacterial community during replenishment after soil fumigation

We compared the responsiveness and sensitivity to soil fumigation of DNA- and RNA-based analyses of a bacterial community. We first established an improved RNA extraction method using DNA as an adsorption competitor, because it is extremely difficult to extract nucleic acids from clay-rich volcanic ash soil (Andisol), which adsorbs nucleic acids. This novel method facilitated RNA extraction from 500 mg of Andisol for molecular analyses. Then we monitored 16S rDNA PCR and 16S rRNA RT-PCR denaturing gradient gel electrophoresis (DGGE) profiles of samples collected from a chloropicrin (CP)-treated field over 2 months. The difference between untreated control and CP-treated plots was detected clearly both in DNA- and RNA-based DGGE profiles after treatment. The temporal changes in DGGE profiles, however, differed between DNA- and RNA-based analyses in CP-treated plots. RNA-based DGGE showed quicker and greater changes in the bacterial community after CP treatment than did DNA-based DGGE, which showed similar trends to RNA-based DGGE but with a time lag. The extent of decrease in the diversity index (H′) and the change in principal response curves was larger in RNA-based analyses. These results indicate that the rDNA PCR-DGGE method also detects DNA of microbes no longer alive after fumigation, and that rRNA provides a more responsive biomarker than rDNA.     

五大连池火山岩土壤发育过程中根际细菌多样性分析

选取五大连池火山群中三座火山为研究对象,采用PCR-DGGEB联用技术研究火山岩土壤发育过程与根际细菌多样性间相关性。结果表明,随火山喷发后时间推移,三座火山南坡有机质和全氮含量先增后稳,北坡土壤有机质和全氮含量先增后减,南北坡土壤p H不断升高。运用Bio-Dap软件分析发现,随时间推移,三座火山南坡细菌丰富度和多样性指数变化趋势为先增后减,北坡为先减后增,南北坡均匀度变化趋势均先增后稳。采用SPSS软件作相关性分析,三座火山南北坡根际细菌多样性指数及丰富度与土壤有机质显著正相关(P0.05)。根际细菌群落多样性受火山岩土壤发育程度影响。