不同提取方法的铁线莲属叶片总DNA提取效果

以铁线莲属植物的叶片为材料,采用CTAB-SDS法、简易CTAB法、改良CTAB法等3种提取方法,比较耗时、纯度、得率、浓度和质量等指标,以期为铁线莲属植物叶片总DNA的提取筛选适宜方法.结果表明,在纯度和浓度方面CTAB-SDS法＞简易CTAB法＞改良CTAB法,在节省提取时间方面改良CTAB法＞CTAB-SDS法＞简易CTAB法.以CTAB-SDS法所提取的9种铁线莲DNA为模板,进行PCR扩增,CTAB-SDS法提取的DNA得率高、质量好,PCR扩增成功率也较高.综合比较,采用CTAB-SDS法进行铁线莲属叶片总DNA提取效果更佳.     

思茅松大配子体DNA的提取及ISSR反应体系的建立

为建立思茅松大配子体ISSR分子标记的反应体系,以思茅松种子样本作为供试材料,比较不同思茅松胚乳DNA提取方法的效果,通过紫外分光度计、DNA琼脂糖电泳和PCR扩增分别予以检测并比较。结果表明SDS法优于CTAB法。同时,通过单因子分析法对ISSR-PCR反应体系中的模板DNA用量,Taq DNA聚合酶用量,Mg2＋浓度,dNTP浓度、引物浓度等主要影响因子进行优化,确立适用于思茅松的ISSR分子标记反应体系。对该体系进行稳定性检验,结果表明该反应体系适合思茅松ISSR-PCR扩增。     

红山茶总DNA提取及ITS-PCR扩增条件优化研究

利用CTAB法对红山茶组植物总DNA进行提取研究,发现用嫩叶可获得纯度较高的DNA,样品在-20℃低温下短期保存对DNA的提取质量无明显影响,同时,对红山茶PCR扩增条件中的各个因素也进行了多梯度的优化研究,建立了引物浓度0.5μmol/L、Mg2 浓度为2.0～2.5mmol/L、dNTP浓度为0.2mmol/L、Taq酶的用量为1.2U、DNA模板用量为50～60ng的最佳ITS-PCR扩增条件。     

白草种子基因组DNA五种提取方法的比较

用5种方法 (传统CTAB法、改良CTAB法、高盐低p H法、改良SDS法、试剂盒法)对白草种子基因组DNA进行提取,分别用紫外分光光度计法、琼脂糖凝胶电泳法、PCR扩增检测提取的DNA的质量。结果表明,传统CTAB法提取的DNA浓度低,含有较多杂质,抑制PCR反应;改良SDS法和改良CTAB法提取的DNA纯度低,不符合检测要求;高盐低pH法和试剂盒法提取的DNA纯度高、完整性好,但试剂盒法成本高、DNA得率低。所以高盐低pH法为5种方法中最经济、效果最好的白草种子DNA提取方法。     

枣叶片DNA的提取以及AFLP反应体系的建立

扩增片断长度多态性(AFLP)实验过程中,DNA提取质量和适合的PCR扩增反应体系是2个关键因素.利用CTAB法对枣树叶片DNA的提取方法进行探讨.结果表明:改良CTAB法抽提到了较高质量的枣基因组DNA,可用于AFLP实验分析;通过对酶切-连接体系和PCR扩增体系中的关键因素实验分析,建立了适合枣AFLP荧光反应体系,并得到清晰的DNA指纹图谱.     

壳斗科5属植物基因组DNA提取方法研究

植物体内所含的蛋白质、酚类、单宁、色素及多糖等物质影响Tag酶的聚合效率,是使PCR 扩增反应失败的主要因素.本研究以壳斗科植物为材料,对CTAB法、SDS法提取的DNA 进行了比较研究,结果表明:CTAB法适合于壳斗科植物基因组DNA的提取,采用这种方法提取的DNA能很好的用于PCR扩增.     

枫香ISSR-PCR扩增条件的优化

对浙江省天台山枫香自然种群的DNA进行提取和ISSR-PCR扩增条件的优化。分析了Mg2+浓度、4×dNTP浓度、BSA浓度、模板DNA用量、引物用量、Taq DNA聚合酶用量等对反应结果的影响。建立了适用于枫香ISSR分析最适宜的反应体系:10μl PCR反应体积中,1×Taq酶配套缓冲液(10 mmol.L-1Tris-HCl,pH值9.0,50 mmol.L-1KCl,0.1%TritonX-100),2.0 mmol.L-1Mg2+,0.15 mmol.L-14×dNTP,2 mg.mL-1BSA,10 ng模板DNA,6 pmol引物,0.9 U TaqDNA聚合酶,引物UBC808的最适退火温度为52.4℃。用此反应体系对天台山枫香自然种群进行了遗传多样性分析,结果显示其多态位点百分率为84.26%,Nei指数为0.2665,Shannon信息指数为0.4044,其遗传多样性处于较高水平。     

米槠叶片基因组DNA的提取及分析

以米槠(Castanopsis carlesii)群落的叶片为材料,采用改良的CTAB法提取米槠基因组DNA,并对获得的基因组DNA进行浓度检测、PCR扩增、琼脂糖凝胶电泳检测、含量测定分析以及扩增结果测序。结果表明:米槠叶片提取的基因组DNA质量和浓度较好,具有良好的完整性,PCR效果好,电泳图谱条带显示清晰,测序结果长度符合要求。利用该方法提取的DNA可用于米槠遗传多样性的分析,可进一步为米槠和格氏栲等其它栲属植物的DNA提取以及其它分子研究提供参考。     

湿地松、加勒比松及其杂交种DNA的提取与微卫星PCR反应体系的优化

文章归纳总结了湿地松、加勒比松及其杂交种湿加松DNA提取的操作流程及其质量检测等方法,并就湿加松的微卫星PCR反应体系的优化问题,进行了一系列的试验,包括Mg2 浓度的梯度试验、不同退火温度和不同循环次数下的扩增试验。试验结果表明,用CTAB法在冰浴下研磨就可以得到质量较好的 DNA用于PCR扩增;PCR的最优反应体系使用20μl的反应体积,2．0 mM镁离子浓度,经30个循环后可扩增出较好的谱带。     

八角基因组DNA的提取方法

采用4种植物基因组DNA提取方法(CTAB法、SDS法、高盐法、试剂盒法)对八角基因组DNA进行提取试验.结果表明,对传统CTAB法稍加改良可以得到较为理想的提取效果,改良的内容包括①在加入CTAB提取缓冲液之前,先用CTAB-free缓冲液抽提1～2次;②将CTAB提取缓冲液中CTAB的用量从2％提高到3％;③提取缓冲液的用量由样品干重的10倍增加到60倍.用改良CTAB法提取的DNA经RAPD-PCR扩增,条带清晰,其质量能够满足分子标记的要求.     

Decomposition and nutrient loss from prunings of different contour hedgerow species in tea plantations in the sloping highlands of Sri Lanka

Contour hedgerows of multipurpose tree species in the sloping tea lands of Sri Lanka are expected to reduce soil erosion and also add significant amounts of plant nutrients to the soil via periodic prunings. The objective of this experiment was to characterize the biomass decomposition pattern and quantify the amount of nutrients added through prunings of six tree species (Calliandra calothyrsus, Senna spectabilis, Euphatorium innulifolium, Flemingia congesta, Gliricidia sepium and Tithonia diversifolia) currently being used in hedgerows associated with tea. Withered leaf and stem prunings (50 g) were enclosed in 2-mm litter bags, placed at 5-cm depth and retrieved after one, three, six, nine and 12 weeks. Loss of initial dry weight, N, P and K was measured. Single exponential decay function adequately described both dry weight and nutrient loss. Tree species differed significantly in their rate of breakdown with decomposition constants (k) varying from 0.0299 to 0.2006 week⁻¹ for leaves and from 0.0225 to 0.0633 week⁻¹ for stems. Gliricidia showed the highest k for leaves with the rest in the following descending order: Senna > Tithonia ≥ Euphatorium > Calliandra > Flemingia. A similar pattern was observed for loss of all nutrients with Calliandra and Flemingia always having lower k values than the rest. Although N immobilization was not observed, immobilization of P and K was observed during the first week of incubation in some species, particularly in stem prunings. Annual biomass of prunings differed significantly between tree species in the following descending order: Calliandra > Senna > Flemingia > Tithonia > Gliricidia > Euphatorium. Calliandra added the greatest amount of nutrients annually to the soil with Euphatorium adding the least. Calliandra prunings provided the annual total K requirement and 49% of the N requirement of mature tea. However, none of the species provided more than 5% of the P requirement. It is concluded that among the tree species tested, Calliandra and Flemingia are the most suitable for contour hedgerows in tea plantations of this agroclimatic region because of their higher soil nutrient enrichment capacity and slower decomposition rates which would minimize leaching losses. This revised version was published online in June 2006 with corrections to the Cover Date.     

Effects of Seed Weight and Seed Type on Early Seedling Growth of Pinus sylvestris under Harsh and Optimal Conditions

The effects of seed weight and seed type on seedling growth of Pinus sylvestris (L.) were studied by seeding individually weighed orchard and stand seed in different mixtures under harsh (direct seeding in field) and optimal (seeding in nursery) conditions. In the nursery experiment an increase in the seed weight from 3 to 7 mg increased the seedling height by 10-27% and total weight by 27-113%, and decreased the height/diameter ratio by 5-6% after 2 yrs. With elimination of seed weight effects, orchard seedlings were 2% taller than stand seedlings in year 2. Without elimination of seed weight effects, orchard seedlings were 7-13% taller. In the field experiment an increase in the seed weight from 3 to 7 mg increased seedling height by 18-65%, stem volume by 81-274% and the number of top-buds by 23-34% in year 5. After elimination of seed weight effects, orchard seedlings were 7-13% taller than stand seedlings and without elimination of seed weight effects 20-21% taller after 5 yrs. Even after elimination of both seed weight and genetic effects orchard seedlings were 3-9% larger than stand seedlings in the field experiment. In conclusion, the influence of seed weight and seed type on growth traits and slenderness is highly significant and the influence seems to be greater in harsh conditions.     

Modelling of adequate pretwist for obtaining straight timber

Abstract

Wood in general and wooden studs in particular are often distorted owing to uneven shrinkage during the drying process in the sawmill. Twist is often the most detrimental of all types of distortion, and it is caused by spiral grain in combination with variations in moisture content. For sawmills, the objective is to produce dried, straight boards, and one method of dealing with boards with excessive spiral grain is to sort them out and then dry them in a pretwisted position to obtain straight boards after drying. A model using the finite element (FE) method for the simulation of drying twist distortions was first calibrated against laboratory experiments in which boards were dried with and without restraints and pretwists. After the calibration, the FE results were compared with industrial test results for boards that were dried without restraints or with restraints with zero pretwist, i.e. straight restraints. The FE model used an elastic–ideally plastic material model to obtain permanent deformations. The calibration was to set the yield stresses so that there was a good match between FE results and results from the laboratory experiments. The comparison between the industrial test results and the FE results showed that the FE model is capable of realistic simulations of drying boards with and without restraints and presumably also pretwists.     

Potential to harness superior nutritional qualities of exotic baobabs if local adaptation can be conferred through grafting

Baobab leaves form an important part of the local diet in Sahel countries and elsewhere in Africa. Existing leaf nutritional data and agroforestry performance information are based solely on Adansonia digitata L., the baobab of continental Africa. The introduction potential of Adansonia species from the center of diversity in Madagascar and from Australia remains untapped. To assess this potential, the mineral contents and B₁ and B₂ vitamin levels of dried baobab leaves were determined for five-year old trees of A. digitata, A. gibbosa (A. Cunn.) Guymer ex D. Baum, A. rubrostipa Jum. & H. Perrier (syn. A. fony Baill.), A. perrieri Capuron and A. za Baill. grown in an introduction trial in Mali. Nutritional data were evaluated against survival and vigor to identify promising germplasm. Leaf vitamin and crude protein contents were highest in the Madagascar species, especially A. rubrostipa (B₁ 88 mg 100 g⁻¹, B₂ 187 mg 100 g⁻¹, protein 20.7% dry weight). However, the local species far outperformed the introductions in survival, tree height, basal diameter and resistance to termites. We suggest grafting as a way of harnessing the vigor of well-adapted local baobab varieties to the superior nutritional profiles of A. rubrostipa and others. Cross-species grafting tests in Adansonia were successful, thus creating new agroforestry possibilities with different scion/rootstock combinations.     

Biogeochemical impacts of clearfelling and reforestation on blanket-peatland streams: II. major ions and dissolved organic carbon

Streams and drains in blanket-peatland forest in western Ireland were sampled weekly over 5 years, 1996–2000, using continuous, depth-proportional passive sampling. Analysis was for pH, alkalinity by Gran titration, anions by IC, metals by ICP, aluminium speciation by loaded-resin exchange, and dissolved organic carbon (DOC) by absorbance at 320 nm.

Effects of felling are identified graphically from two forest drains whose catchments (both about 1 ha) were clearfelled, and partial clearfelling of one larger catchment (somewhat over 1 km²), in summer 1999. Calcium concentrations and alkalinity were increased in the drain- and streamwater from the felled areas. The pH was increased in drains, while in the larger stream, pH range decreased while the mean increased. Phosphorus increased markedly with clearfell, in the absence of recent fertilising, and also increased with fertilising; these results are reported in an accompanying paper. Ammonium-nitrogen concentrations were increased in the two drains with felling, and temporarily decreased in the larger stream. Nitrate increased in some cases. Potassium and manganese concentrations also increased. Concentrations of DOC and organic monomeric aluminium increased gradually, subject to a continuing strong annual cycle. There were no clearfelling effects on concentrations of solphate, suspended solids or inorganic monomeric aluminium. In the two drains (fully clearfelled), concentrations of sodium, chloride and magnesium, and conductivity, were all reduced after felling. Fertilising effects other than for phosphorus were indistinct.

Effects of the combined clearfell–reforestation treatment were testable statistically, using randomised intervention analysis, between two larger streams (1 km²) as a treatment–control comparison. There were statistically significant positive responses in streamwater concentrations of nitrate, potassium, calcium, DOC and aluminium fractions. The implications for management practice depend on whether the loss of nutrients is sustainable for future crop productivity, and whether critical limits and loads of receiving ecosystems are exceeded.     

Effect of Drought on Growth and Mortality of Actively Growing Norway Spruce Container Seedlings Planted in Summer

Survival, root egress and height growth of 13-month-old actively growing Norway spruce ( Picea abies (L.) Karst.) container seedlings exposed to preplanting drying treatments (0, 4, and 8 days) and postplanting drought periods (1, 2, 3, and 4 weeks) were studied in the field between 5 July and 18 August 1999. The mortality of seedlings increased and the height growth and root egress decreased throughout the postplanting drought period. Postplanting performance was also affected by preplanting drying treatments. The results indicate that no risk of excessive mortality and growth restrictions occurs when actively growing seedlings are planted in summer, provided that seedlings are well watered before planting and the drought period does not exceed 3 weeks.     

Rehabilitation of diseased cacao fields in Peru through shade regulation and timing of biocontrol measures

The objective of this case study was to test the effectiveness of biocontrol for the rehabilitation of abandoned, diseased (moniliasis, witches' broom and black pod) cocoa and to develop testable hypotheses regarding the effect of shade and biocontrol timing. Three trial fields differed in their shade regimes. Biocontrol was administered starting either at flowering or pod filling. Disease incidence and yield were compared with cultural control alone. Biocontrol reduced moniliasis from 49% to 25%, independent of the shading regime and application time. Witches' broom was most severe without shade and unaffected by biocontrol. No significant effects on black pod were observed. Although biocontrol increased percentage healthy fruit independent of shading and timing, only the shaded plot with biocontrol from flowering onwards responded with increased yield. Our hypothesis is: 'In the rehabilitation of abandoned cacao, only fields under sustainable shade management and early onset of biocontrol can realise their production potential.' This revised version was published online in June 2006 with corrections to the Cover Date.     

qPCR quantification of Ophiognomonia clavigignenti‐juglandacearum from infected butternut trees under different release treatments

The use of a molecular assay for quantifying conidia of Ophiognomonia clavigignenti‐juglandacearum, the fungal pathogen responsible of butternut canker, was investigated. Purified DNA from conidia collected on glass fibre filters of a passive rain collectors was quantified using a TaqMan real‐time quantitative polymerase chain reaction (qPCR) assay. The qPCR assay could specifically discriminate the target species from all other North American known species of Ophiognomonia, and it was sensitive enough to repeatedly detect one conidium. A linear relationship between numbers of conidia and qPCR C_t values was determined, and used to assess the sporulation of the pathogen under trees that were released to promote their vigour. In total, 977 samples of field‐captured conidia from 49 trees, at two locations, and from two successive growing seasons were analysed. No significant difference of sporulation was observed under control and release treatments. However, our results demonstrated that qPCR assay was reliable for detecting and quantifying O. clavigignenti‐juglandacearum from environmental samples, which will be useful to assess further control methods for this disease.     

The potential of alley cropping in improvement of cultivation systems in the high rainfall areas of Zambia I. Chitemene and Fundikila

Three trials investigating the potential of alley cropping to improve the traditional systems of cultivation, chitemene and fundikila, in the Northern Province of Zambia are described. Flemingia congesta, Tephrosia vogelii, and Sesbania sesban, were grown in association with finger millet, groundnut, cowpea, and maize in various traditional cropping sequences. The indigenous species Tephrosia vogelii and Sesbania sesban were not able to withstand repeated pruning and the long following dry season, and were replaced with Calliandra calothyrsus, and Cassia spectabilis.Over a four year period, there was no benefit by alley cropping with any of the tree species on crop yields, and yields in alley crop treatments even dropped significantly below the control treatments in the fourth year, casting doubt on the potential of alley cropping for sustainable production. There was also no consistent effect on soil chemical characteristics in any of the trials. It was suggested that this lack of beneficial response to alley cropping was due to low tree biomass production low quality of prunings, and an inappropriate cropping sequence. There was no evidence that alley cropping contributed to enhanced nutrient recycling, despite substantial localised pools of soil nutrients, particularly in the chitemene, with which recycling could potentially occur.     

Toxicity of biocide GCSC-BtA on arthropod pests under different temperature conditions

The present research dealt with the toxicity of GCSC-BtA (Germany–China Scientific Cooperation-Bacillus thuringiensis-Abamectin), a new type biocide developed by conjugating a toxin from Bacillus thuringiensis (B.t.) with Abamectin from Streptomyces avermitilis, on arthropod pests under different temperature conditions. The results showed that GCSC-BtA possessed higher toxicity than B.t. crystal, Abamectin or Cypermethrin, with pest mortalities of 97.9, 93.7, 96.4, 82.3 and 96.7% for Tetranychus cinnabarinus (Boisd.) (Acari, Tetranychidae), Frankliniella occidentalis Pergande (Thys., Thripidae), Aphis fabae Scopoli (Hom., Aphididae), Plutella xylostella (L.) (Lep., Plutellidae) and Cameraria ohridella Deschka et Dimić (Lep., Gracillariidae), respectively. Toxicities of GCSC-BtA to T. cinnabarinus and P. xylostella decreased significantly at the older developmental stage with an exception that egg stage had the highest tolerance, in which LC₅₀s of GCSC-BtA were 0.0001, 0.0019 and 0.0708 mg/ml for nymph, adult and egg of T. cinnabarinus, and that 0.0399, 0.2035 and 0.9033 mg/ml for the 2nd instar larvae, 4th instar larvae and egg of P. xylostella, respectively. Also, the biocide was more effective to the lower stage of A. fabae than the higher one with LC₅₀s of 0.0023, 0.0086 and 0.0171 mg/ml against 1st instar nymph, 3rd instar nymph and adult, respectively. In general, toxicity of the new type of biocide was positively related to temperature against all the tested pests, except against T. cinnabarinus, where it was almost similar at the three temperature conditions. GCSC-BtA displayed significantly lower toxicities at 15/20°C than 20/25°C or 25/30°C against F. occidentalis, A. fabae, P. xylostella and C. ohridella. However, no significant difference was found in the toxicities against F. occidentalis, A. fabae and C. ohridella above 20°C, whereas toxicity against P. xylostella increased as temperature raised. Therefore, to achieve higher efficacy in the field, application rate and time should be devised according to these factors.