Status and Advances of Molecular Genetic Improvement of Poplar Species in China

Poplars are among the most important deciduous tree species in China. China is replete with natural resources of poplars. Poplars have a number of good characteristics, including fast growth rate, high yield, many uses, easiness of tissue culture and small gene group that make them well suited as a model system for the application of genetic engineering in forest trees. In the last decade, much progress has been made in genetic improvement of poplar species in China. Modern biotechnology is an important tool for genetic improvement in forest trees, and its applications to genetic improvement in poplars, which covers genetic transformation, gene expression, construction of genetic linkage map, QTLs (quantitative trait loci) identification and molecular assisted selection are reviewed in this paper. At the same time, the existing problems and outlook about the application of modern biotechnology to genetic improvement in forest trees are also discussed.

Cement-bonded particleboard with a mixture of wheat straw and poplar wood

We investigated the hydration behavior and some physical/mechanical properties of cement-bonded particleboard （CBPB） containing particles of wheat straw and poplar wood at various usage ratios and bonded with Portland cement mixed with different levels of inorganic additives. We determined the setting time and compression strength of cement pastes containing different additives and particles, and studied the effects of these additives and particles on thickness swelling, internal bond strength and modulus of rupture of CBPB by using RSM （Response Surface Methodology）. The mathematical model equations （second-order response functions） were derived to optimize properties of CBPB by computer simulation programming. Predicted values were in agreement with experimental values （R2 values of 0.93, 0.96 and 0.96 for TS, IB and MOR, respectively）. RSM can be efficiently applied to model panel properties. The variables can affect the properties of panels. The cement composites with bending strength 〉 12.5 MPa and internal bond strength 〉 0.28 MPa can be made by using wheat straw as a reinforcing material. Straw particle usage up to 11.5% in the mixture satisfies the minimum requirements of International Standard, EN 312 （2003） for IB and MOR. The dose of 4.95% calcium chloride, by weight of cement, can improve mechanical properties of the panels at the minimum requirement of EN 312. By increasing straw content from 0 to 30%, TS was reduced by increasing straw particle usage up to 1.5% and with 5.54% calcium chloride in the mixture, TS satisfied the EN 312 standard.

Isolation and analysis of a TIR-specific promoter from poplar

A 5＇ flanking region of the well-conserved Toll/interleukin-1 receptor domain （TIR）-encoding sequence was isolated from the genomic DNA ofMelampsora magnusiana Wagner resistant clones of hybrid triploid poplars [（Populus tomentosa × P. bolleana） × P. tomentosa]. Sequencing results and alignment analysis show that the obtained TIR-specific promoter （named as PtTIRp01） was 1,732 bp in length; moreover 3＇ region of the PtTIRp01 contains a responds to the 5＇ composition of TIR-NBS type gene PtDRG02, of 747 bp long 5＇ region of TIR-NBS type gene PtDRG02 and its 398 bp complete TIR-encoding sequence, which significantly corindicating that the obtained TIR-specific promoter region consists upstream region of promoter （985 bp）. It was found that the 5＇ region of TIR-NBS type gene PtDRG02 was characterized in the downstream region of the transcriptional start, named as 5＇-untranslated region （5＇ UTR）, consisting of one 93 bp 5＇-untranslation exon, one 213 bp intron and one 441 bp TIR-encoding open reading frame （ORF）. In addition, several putative cis-acting motifs were present in the obtained TIR-specific promoter of PtDRG02, including one TATA box, one GC-rich, one AT-rich, one P-box, one 3-AF1 binding site, two CAAT boxes, two GT-1 motifs, three typical W-boxes, four I-boxes, and one multi-cis-acting fragment （MCF）. The latter contains five types of regulatory elements （E4, G-box, ABRE motif, box 1 and HVA 1 s）, most of which were homologous to the cis-acting regulatory elements involved in the activation of defense genes in plants. Thus, it can be suggested that TIR-specific promoter might be a pathogen-inducible promoter and be necessary for the inducible expression of defense-related genes. Key words Toll/interleukin- 1 receptor domain, promoter, Cis-acting element, poplar

Variation of soil enzyme activity and microbial biomass in poplar plantations of different genotypes and stem spacings

To improve the productivity of poplar planta- tions, a field experiment of split-plot design with four tree spacings and three poplar clones was established, and four soil enzyme activities and microbial biomass were monitored in the trial. Soil enzyme activities, in most cases, were significantly higher in topsoil （0-10 cm） than in lower horizons （10-20 cm）. Soil cellulase, catalase and protease activities during the growing season were higher than during the non-growing season, while invertase activity followed the opposite trend. Soil invertase, cellulase and catalase activities varied by poplar clone but soil protease activity did not. Cellulase and protease activities in the plantation at 5 × 5 m spacing were significantly higher than in the other spacings. The highest catalase activity was recorded at 6 × 6 m spacing. At the same planting density, invertase activity was greater in square spacings than in rectangular spacings. Soil microbial biomass was also significantly affected by seedling spacing and poplar clone. The mean soil MBC was significantly lower in topsoil than in the lower horizon, while MBN showed the opposite pattern. Significantly positive correlations were observed among soil cellulase, protease and catalase activities （p 〈0.01）, whereas soil invertase activity was negatively and significantly correlated with cellulase, protease and catalase activities （p 〈 0.01）. Soil microbial biomass and enzyme activities were not correlated except for a significantly negative correlation between soil MBC and catalase activities. Variations in soil enzyme activity and microbial biomass in different poplar plantations suggest that genotype and planting spacing should be considered when modeling soil nutrient dynamics and managing for long-term site productivity.

Progress and Strategies in Cross Breeding of Poplars in China

The advance in intrasection and intersection cross breeding of poplars in China over the past 50 years is reviewed. Great progress has been made in Sections Leuce and Aigeiros, and satisfactory results of intersection hybridization have been achieved in the crossing between Sections Tacamahaca and Aigeiros. The modes of hybridization include single cross, double cross, triple cross, backcross, etc. It is known that using hybrids as parents to cross with other species or hybrids is an effective and easy way to obtain heterosis. Fast growth, cold and drought tolerance, pest and disease resistance, narrow crowns and rootage, etc. are breeding goals. The conventional artificial crossing is still a major breeding method, and a combination of the conventional artificial crossing with physical radiation and chemical induction can create new triploid individuals that possess higher yield potential. The super clones cultivated have already displayed enormous socioeconomic and ecological benefits in practice. Finally, the problems that investigators have to face at present are discussed as well as some strategies in poplar cross breeding in China.

Variation analysis of growth traits of four poplar clones under different water and fertilizer management

To determine appropriate quantities of water and fertilizer required for early growth of hybrid poplar cuttings,we recorded the growth traits of four clones grown under four factors(irrigation and nitrogen,phosphorus,and potassium fertilizers),each with four levels,using an orthogonal experimental design.A logistic model was used to estimate growth in height.The growth curves for tree height were sigmoid,and the model R^2 values were greater than 0.9,which indicated that the fit was highly significant.ANOVA results for tree height and basal diameter indicated that all sources of variance showed significant differences(p<0.001).The average tree height and basal diameter for all the four clones under the different treatments ranged from 155.39 to 235.04 cm,and from 13.71 to 17.42 mm,respectively.A highly positive correlation between the extreme k value and tree height was observed,suggesting that the k value was an accurate estimation of tree height.For model parameters,the earliest average time point for the onset of the rapid growth period of poplar clones was 131 d,and the highest average increment in tree height during the rapid growth period was 138.78 cm.The highest average tree height for all clones under each factor was 219,210.51,200,and 201 cm when treated with either 1200 mL of water applied every third day,3 g of nitrogen,0 g of phosphorus,or 0 g of potassium,respectively.The most suitable treatment for the early growth of hybrid poplar cuttings,as suggested by the developed logistic model,was 1200 mL of water applied every third day and three applications of 1 g nitrogen(in the form of CH4N2O).

Differences of EST-SSR and genomic-SSR markers in assessing genetic diversity in poplar

We analyzed the genetic differences of 16 poplar clones between genomic-SSR and EST-SSR markers. The statistical results show that the average number of alleles detected by genomic-SSR was 4.1, Shannon＇s index 1.0646, observed heterozygos- ity 0.4427 and expected heterozygosity 0.5523, while for the EST-SSR, the average number of alleles was 2.8, Shannon＇s index 0.6985, observed heterozygosity 0.2330 and expected heterozygosity 0.4684. Cluster analysis indicated that the EST-SSR capacity of genotypic identification was more precise than that of genomic-SSR. These resuks reveal that EST-SSR and genomic-SSR have statistically significant genetic differences in polymorphism detection and genotypic identification. These differences could provide a theoretical basis for the rational use of SSR markers in species diversity and other related research.

Use of amino silane coupling agent to improve physical and mechanical properties of UF-bonded wheat straw(Triticum aestivum L.) poplar wood particleboard

We evaluated the potential use of amino silane coupling agent （SiNH） to improve physical and mechanical properties of UF-bonded wheat straw （Triticurn aestivum L.） poplar wood particleboard. We examined the effects of varied content of silane coupling agent content and ratios of straw to poplar wood particles on particleboard prop- erties. The ratios of straw to poplar wood particles were 100：0, 85：15, 70：30 and 55：45. Silane coupling agent content was tested at three levels, 0, 5 and 10 %. The experimental panels were tested for their mechanical strength, including modulus of elasticity （MOE）, modulus of rupture （MOR）, intemal bonding （IB） and physical properties according to procedures specified in DIN 68763 （Chipboard for special purposes in building construction： concepts, requirements, testing, 1982-03, 1982）. All board properties were improved by the addition of silane cou- pling agent. The use of poplar wood particles had a positive effect on the mechanical properties of wheat straw parti- cleboard but had a negative effect on physical properties （thickness swelling and water absorption）.

Dynamic changes of phenolic compound contents in leaf and bark of poplar during autumn temperature drop

We investigated seasonal dynamics of phenolics substance in leaf and bark of two cultivars of poplar, Populus alba×berolinensis and P. בZhonglin Sanbei 1’, during autumn temperature drop for analyzing the roles of phenolic secondary metabolites in cold resistance. Results show that the contents of condensed tannin and flavonoid in poplar leaf and the flavonoid contents in bark of P. בZhonglin Sanbei 1’ were increased with the decrease of autumn minimum temperature, showing a significantly negative correlation between the contents of soluble phenolic substance in the leaves and changeable temperature. In contrast, lignin content in the poplar leaves is decreased in the process of temperature drop, showing a significantly positive correlation. These results indicate that the variation in phenolic substance has a close correlation with its cold-resistance during the autumn temperature drop.

A recommended rate for application of Chaetomium globosum ND35 fungus fertilizer on poplar plantations in China

Previous studies showed that Chaetomium globosum ND35 fungus fertilizer can improve the microbial community structure and enzyme activities of replanted soil. However, it remains unclear whether can improve the physiological and ecological characteristics of plants under successive rotation. In this study, we investigated the photosynthetic, physiological, and biochemical indexes including photosynthetic parameters, chlorophyll fluorescence, and chlorophyll content of 1-yeax-old poplar seedlings under seven different doses （range from 0 to 1.67 g kg-1） of C. globosum ND35 fungus fertilizer to study the effects of fungus fertilizer on photosynthesis of Poplar. Our results showed that： （1） With increasing application of fungus fertilizer in replanted soil, chlorophyll content of poplar leaves （Chl） increased, while physiological indexes such as electron transport rate （ETR）, net photosynthetic rate （Pn）, quantum efficiency （φ）, nitrate reductase （NR） activity and root vigor initially increased and then declined. Meanwhile, heat dissipation that depended on the xanthophyll cycle declined and nonphotochemical quenching （NPQ） initially increased and then decreased. When the dose of C. globosum ND35 fungus fertilizer was 0.67 g kg-1 （T3） and 1.00 g kg-1 （T4）, excess light energy of photosynthetic apparatus was reduced, and photosynthetic apparatus distributed more light energy to the direction of photochemical reactions, which improved the efficiency of energy use. Plant height and biomass of leaves, stems, and roots were maximum at T3. We conclude that applying appropriate amounts of C. globosum ND35 fungus fertilizer can improve root physiological activity and capacity for use of light by poplar leaves. This can improve the operating states of the photosynthetic apparatus and lead to increased photosynthetic efficiency of poplar leaves and accumulation of dry matter.This suggests a strategy to alleviate the successive rotation obstacle of soil nutrient depletion.

An Improved System for Shoot Regeneration from Stem Explants of Lombardy Poplar (Populus nigra L. var. italica Koehne)

We developed a system for the regeneration of Lombardy poplar (Populus nigra L. var. italica) shoots from internodal stem explants. Using this system, shoots regenerated from 87% of the stem explants placed on Murashige and Skoog (MS) medium supplemented with 0.1 mg/L indole-3-acetic acid and 0.5 mg/L benzylaminopurine without undergoing callus formation. About 80% of the in vitro regenerated shoots developed roots on MS medium supplemented with 0.5 mg/L indole-3-butyric acid and 0.02 mg/L 1-naphthylacetic acid. Well-rooted seven-to eight-week-old regenerated plants could be transferred to soil for further growth and the survival rate of such plants after three weeks was 88%. The protocol presented here is simple and economical because it does not rely on pre-incubation in callus induction medium or repeated subculture in shoot induction medium containing trans-zeatin, an expensive substance. The in vitro regeneration system presented here could be used for evaluation of radiation sensitivity for Lombardy poplar tissues.

The influences of biotic and abiotic factors on the occurrence and severity of poplar canker disease in Qingfeng County, China and the management implications

Landscape pathology is a research approach that can provide validation of the effectiveness of regional controls of forest disease at a landscape scale. In this paper, we analyzed the effects of stand features, management approaches, and geographical locations on poplar canker disease incidence （DI） and disease severity index （DSI） of individual trees at a 10 km x 10 km mesoscale landscape in Qingfeng County, China. DI varied significantly with stand age, tree densities, and the degree of canopy closure. DI in stands younger than 4 years old was significantly lower than that in the stands over 6 years old and reached the highest value at a stand age of 8-10 years. Overall, DI was positively correlated with stand age, stand density, andthe degree of canopy closure. DI was significantly lower in agro-forest stand patches than in other three patch types, i.e. isolated patch, pure stand patch, and mixed stand patch. Poplar plantations distributed around and near to villages exhibited significantly higher DI mainly due to human activities and herbivores. Fragmentation or connectivity in this mesoscale landscape seemed not impact disease occurrence. DSI was not significantly correlated with stand density, but varied significantly with tree varieties and trees ages. DSI was highest in stands of 10-12 year trees for all poplar varieties we studied here. Plantation density and plantation age were thus critical factors in determining DI and DSI. A logistic predictive model of disease occurrence was developed for the study area, considering varieties, age, height, density, canopy cover, stand types, patch types, management status, and stand geographical locations. Our study here shows that adjustment of stand density by thinning at different plantation ages is an effective approach controlling the occurrence canker disease in short-rotation poplar plantations at the landscape scale.

Temporal and spatial changes in Bt toxin expression in Bt-transgenic poplar and insect resistance in field tests

Extensive planting of Bacillus thuringiensis （Bt）-transgenic plants economically benefits society; how-ever, the potential risk they pose is receiving increasing attention. This study used enzyme-linked immunosorbent assay and fluorescence quantitative PCR （RT-PCR） to monitor the temporal and spatial dynamics of the expression of Bt toxic protein in a forest of 6- to 8-year-old trees of transgenic insect-resistant poplar 741 for three consec- utive years. The enrichment, distribution, and degradation of Bt toxic protein and the influence of transgenic poplars on the targeted insect population, Hyphantria cunea, were investigated. The content of CrylAc toxic protein dynamically changed in transgenic poplar. During the annual growth cycle, the content initially increased, then decreased in the long and the short branches of the crown and in the root system, peaking in August. During the study, the protein did not accumulate overtime. The mRNA transcription of gene CrylAc was almost consistent with the level of the protein, but transcription peaked in July. In the transgenic and control forestland, microscale levels of the CrylAc toxic protein were detected from the soil, but increased accumulation was not observed with the planting year of transgenic poplar. Meanwhile, Bt was isolated and detected molecularly from the soil in the experimental forestland. A systematic investigation of the density of H. cunea in the experimental transgenic poplar forest indi- cated that transgenic Pb29 poplar could resist insects to a certain degree. At peak occurrence of the targeted insects, the density of H. cunea in the experimental forest was significantly lower than in the nontransgenic poplar forest.

Identification of differentially expressed proteins in poplar leaves in-duced by Marssonina brunnea f. sp. Multigermtubi

Black spot disease in poplar is a disease of the leaf caused by fungus. The major pathogen is Marssonina brunnea f. sp. multigermtubi. To date, little is known about the molecular mechanism of poplar （M. brunnea） interaction. In order to identify the proteins related to disease resistance and understand its molecular basis, the clone ＂NL895＂ （P. euramericana CL＂NL895＂）, which is highly resistant to M. brunnea f. sp. multigermtubi, was used in this study. We used two-dimensional gel electrophoresis （2-DE） and mass spectrometry （MS） to identify the proteins in poplar leaves that were differentially expressed in response to black spot disease pathogen, M. brunnea f. sp. multigermtubi. Proteins extracted from poplar leaves at 0, 12, 24, 48, and 72 h after pathogen-inoculation were separated by 2-DE, About 500 reproducible protein spots were detected, of which 40 protein spots displayed differential expression in levels and were subjected to Matrix assisted laser desorption/ionization time of flight mass spectrometry （MALDI-TOF MS） followed by database searching. According to the function, the identified proteins were sorted into five categories, that is, protein synthesis, metabolism, defense response and unclassified proteins.

Integrated Control Technique of Poplar Diseases in China

The main disease species on poplar are introduced in this paper. The main content of the integrated control technique on poplar diseases is summarized:(1) Forecast technique based on the initial disease and provention factors. (2) Control index is established, based on the relationship among the disease index, height of tree, diameter grade and the loss rate of volume. (3) Five fine varieties are selected according to three integrated indexes of host, such as resistance, volume growth and form ratio; They are 613 (Poulus alba ×P. berolinensis), A15 (P. xiaohei × P. euramericana CV. Polska-15A), A98 (P.xiaohei×(P. simonii×P. nigra) CV. A98), A102 (P. xiaohei) × (P. simonee × P. nigra) CV. A102), L2 (P. simonii × P nigra var. italica) in northeast Chnia. (4) Sivicultural control is known as the main protection measures, combined with chemical control and biocontrol,based on the disease forecast and control index by tracing the whole process of forest production.

Degradation of atrazine and changes in soil biological indices throughout dendroremediation using poplars

Dendroremediation,a novel strategy for cleaning up contaminants from the environment by using special woody plant species and microbes,is a popular phytoremediation approach.It is a cost-effective and environmentally friendly method that is being increasingly adopted.Poplar species(Populus L.)have been suggested for use in remediation because of their characteristics of high biomass production,fast growth,and potential for removing pollutants.Our environment is often contaminated by toxic substances produced by human activities,and remediation of contamination is therefore a global issue.Atrazine is one of the most widely used herbicides in China.In the present study,‘I-69/55’poplar(P.deltoides cv.‘I-69/55’)and hybrid poplar(Populus deltoides×nigra,DN34)grown in pots were assayed for their ability to remediate atrazine-contaminated soils.The degradation dynamics of atrazine were assessed with a high-performance liquid chromatography system using materials from a greenhouse with different rhizosphere environments.We studied the dynamic variation in microbes and microbial biomass carbon to elucidate the rhizosphere effects and mechanism of remediation of atrazine-contaminated soil by poplar.Our four treatments included a control with uncontaminated soil without a tree,atrazine pollution without a tree,atrazine pollution with‘I-69/55’poplar,and atrazine pollution with hybrid poplar.Hybrid poplar showed potential for remediation of atrazine-contaminated soil,and the degradation of atrazine in the rhizosphere was faster than that in non-rhizosphere soil.Atrazine significantly inhibited bacteria growth in nonrhizosphere soil.The high concentration of bacteria in the hybrid poplar rhizosphere might be key to atrazine degradation.Trends of change among fungi and actinomyces did not correspond to trends in atrazine degradation throughout the trial period.Further research is needed to predict the effects of atrazine on cultivable microorganisms in various soils.Atrazine had significant inhibitory effects on microbial biomass carbon in non-rhizosphere soil,and the rhizosphere environment of poplars enhanced the recovery of microbial biomass carbon.The potential for hybrid poplar as a dendroremediation material needs further study.

THEORY AND APPLICATION OF CUTTAGE SEEDLING AND AFFORESTATION ON POPLAR

First, a theoretical model of density of cuttage seedlings on poplar was evaluated by using mathematical tools based on anatomy and morphology of photosynthesis organ, then,according to this, an afforestation density of the trces was put forward, finally, the importance,scope and method of the theoretic application wcre described through a concretc calculation. The result showed that these density models had not only rigorous theory base, including biology and mathematics, but also wide applying prospect. And at the same timc, the model parameters could be gotten easily in forestry practice.

Impact of spacing and rotation length on nutrient budgets of poplar plantations for pulpwood

The above-ground biomass and nutrient accumulation by poplar plantations were evaluated for pulpwood production ]n China, Experimental treatments applied in a split-plot design included four planting densities (1111, 833, 625 and 500 stems hm-2), three rotation lengths (4a, 5a and 6a) and three poplar clones (1-69.NL-80351 and 1-72). The highest biomass was achieved in the highest stocked stand (1111 stem hm-2’) at 6 of rotation age for both clone 1-69 and clone 1-72, which is about two times that in the stands of 500 stems hm-2 at 4 ofrotation age. However, the highest occurred in the stand of 833 stems-hm-2 at 6-year rotation for NL-80351. Ranking of the plantation biomass production by component was stem > branches > foliage> stem--bark and the production of the Support components of the plantation was 10-fold that of the productive component, i.e., foliage. Thepattern of accumulation of nutrients by the plantations was similar to the biomass. Nutrient accumulation in theplantations was in the order of Ca>N>K>Mg>P, but some differences existed in annual nutrient accumulationrates for four planting densities and three poplar clones, The mean annual accumulation of N and P in the plantations was 13.2 and 2.8 kg hm-2 in stem, 12.1 and 1 .9 kg hm 2 in branch, and 98.5 and 9.5 kghm’ in foliage. Themean Ca, K and Mg accumulations were 28.2, 1 8.5 and 2.9 kg’hm-2 a-1, 26,9. 11,0 and 2.3 kg-hm-2-a-1 in branch.and 116.5, 81 .3 and 16, 1 kg hm-2 a-1 in foliage, respectively. Biomass utilization standards markedly affected theexport of nutrients from the site, Whole tree utilization yields the most biomass and removes the most nutrients,Removal of stem with 2 10-cm diameter exports about half of the biomass, but N and nutrients removals are only23% and 28% of the total, respectively, Removal of the entire stem provides about tWo--thirds of the total biomassand removes 31,1% total N and 37.5 % total nutrients respectively. Including the branches in the removal increases biomass yield to 92% of the total, and nutrient removal is about 68% of the total,

Photosynthetic response of poplar leaves at different developmental phases to environmental factors

With economic incentives and interests in fast-growing poplar trees for short-rotation production of fiber and veneer, many new poplar hybrids have been bred and planted in China, but how to match the new poplar clones to suitable sites and maintain their higher growth rates is still not very clear. In this study, the photosynthetic response of poplar leaves at various developmental stages during two seasons (summer and autumn) was explored and mechanistic models for the photosynthesis of poplar leaves at different developmental phases in response photosynthetic active radiation (PAR), temperature, and relatively humidity were established using the optimization software package 1st Opt. Mature poplar leaves in autumn had significantly higher photosynthetic capacity than leaves at other stages and seasons. Based on the models established for poplar leaves at different phases, the main limiting factors for photosynthesis at the research site were high PAR and temperature in the summer and low PAR in the autumn. Our results highlight the importance of selecting suitable sites, pruning and stand density control during the plantation development to maintain higher photosynthetic rates of poplar trees and to establish optimum cultivation patterns for various utilization of poplar plantations.

The use of the white poplar(Populus alba L.)biomass as fuel

We determined the calorific value of white poplar（Populus alba L.） woody biomass to use it as firewood. The value of 19.133 MJ kg-1obtained experimentally shows that the white poplar can be quite successfully used as firewood. Being of a lower quality in comparison with usual beech firewood, the white poplar has similar calorific value. The white poplar has a calorific density of 30.7 % lower than that of current firewood. That is why the price of this firewood from white poplar is lower accordingly. Also, the prognosis of calorific value on the basis of the main chemical elements, being very close to the experimental value（＋2.6 %）, indicates an appropriate value can be achieved to be used for investigation with the chemical element analysis.