Note: Thermal analysis of the long line source electron gun.

We performed thermal analysis for our previously reported [M. Iqbal, K. Masood, M. Rafiq, M. A. Chaudhry, and F. Aleem, Rev. Sci. Instrum. 74, 4616 (2003)], long linear filament electron gun assembly using ANSYS software. The source was set under a thermal load of 3000 °C, to evaluate temperature distribution, thermal strain, and heat flux at various components of the gun. We calculated the maximum heat flux (9.0 W/mm(2)) that produced a thermal strain of 0.05 at the focusing electrodes. However, the minimum value of the heat flux (0.3 W/mm(2)) was at the anode electrodes which correspond to a negligible thermal strain. The gun was validated experimentally showing a uniform cross section of the beam at the molybdenum work plate comparable to the size of the filament. Our experimental and theoretical results are in agreement. The gun had been in continuous operation for several hours at high temperatures without any thermal run-out.

Kinetics studies on the lignan class of natural compounds that inhibits alpha-chymotrypsin.

The mechanism of inhibition of the alpha-chymotrypsin enzyme by two lignans of the fused bistetrahydrofuran series, epiexcelsin (1) and 5'-demethoxyepiexcelsin (2), which were isolated from the Commiphora mukul Engl., was investigated. Lineweaver-Burk and Dixon plots and their secondary replots showed that these compounds were noncompetitive inhibitors of the enzyme. K(i) values for 1 and 2 were found to be 22.29 +/- 0.015 and 336.30 +/- 0.053 microM, respectively.

Fluorescence-based sequencing of double-stranded DNA by hexamer string priming.

A fluorescence-based, T7 (Sequenase) dye terminator method for sequencing double-stranded DNA using strings of three contiguous hexamers as primers and single-stranded binding protein is described. In this method, the circular, supercoiled DNA vector pUC19 is first linearized with a restriction enzyme to create a sequenceable template. Sequencing is then accomplished using three cycles of "denaturation," annealing, and extension/termination. Twenty-two of 33 hexamer strings tested in a controlled study produced acceptable sequence, with read lengths varying from the mid 300s to the low 400s and a base-calling accuracy of at least 97%. To test its potential utility in directed DNA sequencing, the protocol was then used to completely sequence both strands of pUC19. For this test project, a total of 28 hexamer strings was used with an overall successful priming rate of 75%. The current protocol appears to be sufficiently robust to be used in the finishing phase of a shotgun sequencing project and is amenable to semiautomation. Prospects for using the protocol for full-scale directed sequencing as well as for full automation are discussed.

High-quality automated DNA sequencing primed with hexamer strings.

The finishing phase of genome sequencing projects is expensive, in part, because of the cost of de novo synthesis of custom primers and the management burden associated with obtaining and using them for primer walking. One approach to reduce these high costs is the use of a presynthesized library of short oligonucleotides (8-10 bases) rather than long primers. The use of such a library eliminates the need for custom synthesis of oligonucleotides, providing the convenience of priming from any site by combining two to three short oligonucleotides to form a string with the required specificity. The first practical implementation of this strategy presented a robust protocol for using hexamer strings with radioisotopic labelling. Whereas versions of this technique have subsequently been implemented on fluorescent sequencers we felt that there was a need to develop and extensively test a protocol that consistently gave read lengths comparable to dye-terminator sequencing with longer primers. We have developed a new two-cycle fluorescent Sequenase terminator procedure for using hexamer strings. We tested this procedure using a set of 32 different 3 hexamer primer strings, each known to be functional to some degree in radioisotopic sequencing, on single-stranded M13mp18 template and ABI 373 DNA sequencers. The overall success rate of priming with these hexamer primer strings is 97% with the failure of only one string. In this case, the corresponding 18-mer primer also failed to produce usable sequence from M13mp18 template. The average read length from reactions successfully primed with the 31 different hexamer strings was 461 bases with > 99% base-calling accuracy. The current protocol is robust enough to be used in virtually any situation where primer walking on single-stranded templates is used. The success rate and read lengths make it universally applicable to the sequencing of single-stranded templates on automated sequencers. It is also amenable to automation.

A molecular marker based linkage map of Vitis.

Genetic linkage maps of Vitis (2n = 38) have been constructed from a single interspecific hybrid grape population (60 seedlings) of 'Cayuga White' X 'Aurore'. The maps were primarily based on 422 RAPD markers but also included 16 RFLP and isozyme markers. These maps had an average distance of 6.1 cM between markers and were developed using a double-pseudotestcross strategy. The 'Cayuga White' map had 214 markers covering 1196 cM and that of 'Aurore' spanned over 1477 cM with 225 markers. The 'Cayuga White' map consisted of 20 linkage groups, whereas 22 linkage groups comprised the 'Aurore' map. The number of groups reduced to 19, as in some instances two or more groups from one parent showed homology with a single group from the other parent on the basis of markers heterozygous in both parents. Each linkage group ranged in size from 14 to 135 cM in 'Aurore' and from 14 to 124 cM in 'Cayuga White'. These maps provide enough coverage of the genome to allow quantitative trait locus analysis and map-based gene cloning.

Nuclear DNA content of Vitis species, cultivars, and other genera of the Vitaceae.

The nuclear DNA content was analyzed in Vitis species, hybrid cultivars, and genera of the Vitaceae using flow cytometry. Significant variation was found among Vitis species, hybrids, and other genera of the Vitaceae (Ampelopsis and Parthenocissus). DNA content was estimated to range from 0.98 to 1.05 pg/2C within V. labrusca (ns) and 0.86 to 1.00 pg/2C within V. vinifera (ns). Genotypes from Vitis and Parthenocissus were similar in nuclear DNA content (approximately 1.00 pg/2C) whereas they differed significantly from Ampelopsis (1.39 pg/2C). No correlation between DNA content and the center of origin of genotypes of the Vitaceae was noted. Based on the present study, the Vitis genome size is 475 Mbp, 96% of which is non-coding. Knowledge of DNA content is useful in order to understand the complexity of the Vitis genome and to establish a relationship between the genetic and physical map for map-based cloning.

Forest understory biomass heterogeneity is "moisture complex" or associated litter the cause?

Understory biomass heterogeneity underPlatanus occidentalis L.,Quercus alba L.,Celtis occidentalis Pursh., andUlmus americana L. was studied. Soil pH, Ca, Mg, Mn, Cu, NH4 nitrogen, total N, and soil moisture were significantly different under all dominant species. The pattern of soil moisture levels under different tree species was not consistent from one sampling period to the next. The aboveground biomass of understory vegetation also varied significantly under different tree species and was not correlated with variation in any of the soil properties. Maximum understory biomass gain (340%) from May to September occurred under oak trees, where soil moisture and most nutrients were the lowest. On the other hand, sycamore and hackberry had continuous release of allelochemics and the smallest understory biomass gain (103%) during the growing season, even though soil under both species had more moisture and nutrients than the soils under white oak. Eliminating all the factors studied leads to the conclusion that organic substances released in the immediate environment of dominant trees and their litter influenced soils and associated herb growth. In each dominant niche system, understory species may have to develop their own system ("microniche") to minimize the stresses of competition and allelochemics.

Allelopathy in saline agricultural land: Vegetation successional changes and patch dynamics.

In reclamation fields of salt-affected wasteland, five plant communities colonized the undisturbed land, represented byCynodon dactylon, Desmostachya bipinnata, Prosopis juliflora, Sporobolus arabicus, andSuaeda fruticosa. Kallar grass (Leptochloa fusca), a highly salt tolerant plant when cultivated, shared dominance withCynodon, Desmostachya, andSporobolus in 15-month-old fields, whereasPolypogon was the only dominant species in 30-month-old kallar grass fields. Through successional stages, soil pH, salinity, sodicity, and Na, K, Ca + Mg significantly decreased due to leaching. Electrical conductivity successively changed from 13.0 to 3.0 to 1.0, while soil total nitrogen, NH4 nitrogen, NO3 nitrogen and available P significantly increased. In high-density kallar grass fields, six weed species appeared only in well-defined patches and radially eliminated or reduced kallar grass growth. Many soil factors, such as pH, EC, NH4 nitrogen, NO3 nitrogen and available P analyzed in patch vegetation soils, were mostly either comparable or significantly better than those of surrounding kallar grass fields. On the other hand, aqueous extracts of all six invading species and kallar grass significantly reduced kallar grass seed germination to varying degrees. Further, decaying leaf powder of allelopathically suspected species significantly reduced kallar grass biomass, which varied from species to species and in most cases corresponded with field data of kallar grass in patch vegetation. It should be strongly pointed out that allelopathic behavior discussed in patch dynamics was in areas where soil saline-sodic conditions had improved greatly (e.g., EC = from 13.0 to only 1.0) due to kallar grass plantation. Further,Suaeda appeared to be a poor competitor when soil conditions improved for other species as well, and it could not capitalize on its evolutionary strategic trait of performing well in saline-sodic conditions. To our knowledge, this is the first report indicating that allelopathy may be a factor in determining growth and distribution of plants in saline or sodic soils.

Allelopathy in agroecosystems: Wheat phytotoxicity and its possible roles in crop rotation.

The germination rates of cotton and wheat seeds were significantly affected by various extracts of wheat mulch and soils collected from the wheat field. This toxicity was even more pronounced against seedling growth. Five allelochemics: ferulic,p-coumaric,p-OH benzoic, syringic, and vanillic acids, were identified from the wheat mulch and its associated soil. Quantitatively, ferulic acid was found at higher concentrations thanp-coumaric acid in the soil. Various concentrations of ferulic andp-coumaric acids were toxic to the growth of radish in a bioassay. The functional aspects of allelochemic transfer from decaying residue to soil and the subsequent microbial degradation within agroecosystems are discussed, particularly as they relate to wheat crop rotation, with wheat and cotton, in Pakistan.

Effects of pine-produced chemicals on selected understory species in aPinus ponderosa community.

Pinus ponderosa accounted for more than 98% of all tree and shrub stratum stems in a climax community with low herb coverage and aboveground biomass, 35% and 60 g/m(2), respectively. Because of our previous report that nitrification and nitrifying bacteria in the same community were allelopathically inhibited, we speculated that the pine-produced allelochemics might also directly influence the development and growth of the herb stratum. In most cases decaying needles, needle leachate, and field soils significantly reduced germination and radicle growth ofAndropogon gerardii andA. scoparius, pine-associated herbaceous species. Additionally, growth ofAndropogon scoparius seedling radicles was reduced 28-56% by pine needle extracts, 33% by pine bark extracts, and 67% by soil hydrolysate extracts.Andropogon seed germination was reduced 20-25% by pine needles and soil. Phytotoxins identified in various plant parts and associated soils were caffeic acid, chlorogenic acid, quercetin, and condensed tannins. Pine needle water and soil hydrolysate extracts were most inhibitory to the radicle growth of the test species. Thus it appears that the limited growth of the herbaceous stratum in the pine community may be accounted for, in part, by allelopathy. Such allelopathic interactions may have an adaptive ecological significance in various forest and other plant communities.

Accelerated soil mineralization, nitrification, and revegetation of abandoned fields due to the removal of crop-soil phytotoxicity.

In an abandoned corn field, clear-cutting of crop vegetation increased the productivity, species richness, and nonannuals in the following years after abandonment, as compared to the control plots from which crop vegetation was not removed. The increase in plant growth was apparently due to the elimination of allelopathic chemicals from the soil, which normally are released from the standing crop. Removal of vegetation also increased the soil mineralization of Ca(2+), Mg(2+), K(+), NH4 (+) and NO3 (-)-N. This situation encouraged species having higher mineral requirements to rapidly invade the fields in the successive years. Clear-cutting also increased the nitrification process by removing the inhibitors of nitrification. The number ofNitrosomonas was always significantly higher in the harvested plots as compared to unharvested plots. Phenolic phytotoxins were isolated from the crop residue and soil. Further, these phytotoxins were significantly higher in the unharvested crops as compared to clear-cut plots, in most samples. Whatever the direct or indirect additional explanation for increased biomass, nonannuals and richness in successive years, it is clear that the removal of standing crop has a definite influence.