Seasonal Variation in the Ultrastructure of the Cambium in New Zealand Grown Pinus radiata D. Don

Examination of Pinus radiata cambium from trees growing in thecentral North Island of New Zealand has revealed that many ofthe structural changes occurring in other tree species at theonset or cessation of cambial activity are not found in thisspecies. The winter cambium bears a closer resemblance to thesummer cambium than it does to the winter cambium of any otherangiosperm or gymnosperm described in the literature. In particularthere is little change in vacuolar structure, endoplasmic reticulumform, and dictyosomal activity during the year. The only changeswhich take place involve a slight increase in vacuole volume,storage of starch in vacuoles, and a decrease in numbers ofspherosomes during the summer. These observations confirm that,while the degree of cambial activity is reduced in the winter,complete dormancy is absent.     

Evaluation of AFLP for genetic mapping in Pinus radiata D. Don

Efficient construction of reasonable density genetic linkage maps is an essential component of QTL detection programmes. The AFLP technique has been used to produce genetic linkage maps in a range of species. We have developed protocols to generate reproducible AFLP profiles in Pinus radiata and have evaluated the inheritance and informativeness of AFLP markers in this important timber species. The large genome size of P. radiata necessitated increased levels of selection at both the pre-amplification and selective amplification steps of the AFLP protocol to generate reproducible AFLP profiles. Once optimised ca. 41.3 scorable AFLP bands were resolvable through denaturing gels, of which 48.4% were polymorphic in a screen of eight unrelated trees. This level of polymorphism is ca. three times higher than with RAPD markers. The total number of bands and the number of polymorphismic bands per PCR were ca. halved when AFLPs were electrophoresed on non-denaturing gels and stained with ethidium bromide. Using the protocols developed, AFLP is an efficient method for generating the DNA markers required for genetic linkage map construction in P. radiata. This revised version was published online in June 2006 with corrections to the Cover Date.     

An allele responsible for seedling death in Pinus radiata D. Don

 When inbred, most outcrossing species show high mortality, manifested at several life stages. The occurrence of homozygotes for deleterious or lethal alleles is believed to be responsible. Here, we report the identification of an allele responsible for the death of selfed Pinus radiata D. Don seedlings in their first month after germination. Among 291 S₁ seedlings of plus-tree 850.55, 76 died within 1 month of emergence. Their death appears to be caused by a single recessive lethal allele, SDPr (seedling death in Pinus radiata). SDPr is located in a linkage group with 28 RAPD markers, the closest of which is ai05800a. Of the 76 seedlings that died, megagametophytes of 73 could be genotyped. Of these, 71 had the null (no band) allele of ai05800a; only two had the band allele. Of the 190 surviving S₁ diploids that were genotyped, only two individuals were homozygous for the null allele of ai05800a. By two different methods, the map distance between SDPr and ai05800a was estimated to be between 1.0 and 2.7 cM respectively. The frequency of band and null alleles in the combined population of dead and surviving seedlings and in un-sown seeds shows no evidence of selection at this locus prior to germination. Received: 30 September 1997 / Accepted: 29 October 1997     

Competitive processes in a monoculture of Pinus radiata D. Don

Summary The changes between 9 and 40 years of age of the frequency distribution of tree diameters were studied in an experiment in a plantation of Pinus radiata D. Don in South Australia in which the spacing between trees at planting varied. Empirical functions were developed to relate tree diameter to growth rate in diameter, variance of growth rate in diameter and mortality rate. These functions were used in solving a forward diffusion equation to predict future diameter distributions. The contributions from each of the terms in the diffusion equation were examined and it was found that the drift (growth rate in diameter) term made the greatest contribution to the solution, whilst the diffusion (variance in growth rate) term had neglible effect. This implied that competition between individual trees for light was the dominant competitive process operating in this plantation, rather than below ground competition for soil resources. The ramifications of this finding to forest growth modelling are discussed.     

The classification and frequency of ectotrophic mycorrhizas onPinus radiata D. Don. in New Zealand

Summary Eight subtypes and thirteen sub-subtypes of ectotrophic mycorrhizas were identified in New Zealand onPinus radiata. Subtype B occurred most frequently. The maximum occurrence of ectotrophic mycorrhizas was in New Zealand conditions at the end of winter and beginning of spring.     

Signatures of adaptation and genetic structure among the mainland populations of Pinus radiata (D. Don) inferred from SNP loci

Insights into the relative contributions of locus specific and genome-wide effects on population genetic diversity can be gained through separation of their resulting genetic signals. Here we explore patterns of adaptive and neutral genetic diversity in the disjunct natural populations of Pinus radiata (D. Don) from mainland California. A first-generation common garden of 447 individuals revealed significant differentiation of wood phenotypes among populations (P _ST), possibly reflecting local adaptation in response to environment. We subsequently screened all trees for genetic diversity at 149 candidate gene single nucleotide polymorphism (SNP) loci for signatures of adaptation. Ten loci were identified as being possible targets of diversifying selection following F _ST outlier tests. Multivariate canonical correlation performed on a data set of 444 individuals identified significant covariance between environment, adaptive phenotypes and outlier SNP diversity, lending support to the case for local adaptation suggested from F _ST and P _ST tests. Covariation among discrete sets of outlier SNPs and adaptive phenotypes (inferred from multivariate loadings) with environment are supported by existing studies of candidate gene function and genotype–phenotype association. Canonical analyses failed to detect significant correlations between environment and 139 non-outlier SNP loci, which were applied to estimate neutral patterns of genetic differentiation among populations (F _ST 4.3 %). Using this data set, significant hierarchical structure was detected, indicating three populations on the mainland. The hierarchical relationships based on neutral SNP markers (and SSR) were in contrast with those inferred from putatively adaptive loci, potentially highlighting the independent action of selection and demography in shaping genetic structure in this species.     

Accession and cycling of elements in a coastal stand ofPinus radiata D. Don in New Zealand

Summary The accession and cycling of elements in a 14-year-old coastal stand ofPinus radiata D. Don was measured for one year. The element contents (mg m^–2 year^–1) of bulk precipitation and throughfall respectively were: NO₃–N 41, 12; NH₄–N 133, 154; organic-N 157, 396; Na 4420, 9700; K 387, 2900; Ca 351, 701; Mg 486, 1320. Of the increase in element content of rainwater beneath the forest canopy 20% (NH₄–N), 70% (organic-N), 3% (Na), 90% (K), 20% (Ca) and 30% (Mg) was attributed to leaching; the remainder to washing of aerosols filtered from the atmosphere by the vegetation. The canopy absorbed approximately 40 mg m^–2 year^–1 of NO₃–N. Litterfall was the major pathway for the above-ground biogeochemical cycle of N (93%), Ca (96%) and Mg (74%), and leaching was the major (73%) pathway for K.     

Stable transformation and regeneration of transgenic plants of Pinus radiata D. Don

A biolistic particle delivery system was used to genetically transform embryogenic tissue of Pinus radiata. The introduced DNA contained a uidA reporter gene under the control of either the tandem CaMV 35S or the artificial Emu promoter, and the npt II selectable marker controlled by the CaMV 35S promoter. The average number of stable, geneticin-resistant lines recovered was 0.5 per 200 mg fresh weight bombarded tissue. Expression of the uidA reporter gene was detected histochemically and fluorimetrically in transformed embryogenic tissue and in derived mature somatic embryos and regenerated plants. The integration of uidA and npt II genes into the Pinus radiata genome was demonstrated using PCR amplification of the inserts and Southern hybridisation analysis. The expression of both genes in transformed tissue was confirmed by Northern hybridisation analysis. More than 150 transgenic Pinus radiata plants were produced from 20 independent transformation experiments with four different embryogenic clones. Received: 9 May 1997 / Revision received: 18 September 1997 / Accepted: 18 October 1997     

Growth and development of Pinus radiata D. Don: the effect of light quality

Abstract. Plants of Pinus radiata of two physiological ages, juvenile (seedlings and micropropagated plantlets) and adult (rooted cuttings from mature trees), were grown under lighting fromthree combinations of metal halide (MH) and tungsten halogen (TH) lamps for up to 10 months in controlled environment rooms. The three lamp combinations, MH alone, 50: 50 MH: TH and 25:75 MH: TH by wattage, produced red: farred ratios of 4.59, 1.51, and 1.15, respectively. Photosynthetic photon flux density was 700 μmol m⁻² s⁻¹. An increase in proportion of TH lamps markedly increased shoot elongation and internode length, decreased numbers of fascicles per unit stem length and increased the proportion of stem weight in both juvenile and adult material. In addition, in adult material, it increased the number of fascicle initials and expanded fascicles per growth flush, reduced the duration of the 'rest' phase between growth flushes, accelerated the rate of elongation growth during each flush, and increased apical dominance. Tracheid length, but not diameter or wall thickness, was significantly affected by light quality and found to be associated with longer internodes. Any treatment effects on needle weight or length, stem diameter or root weight were non-significant or very small. Different clones from either the juvenile micro-propagated material or the mature rooted cuttings each showed similar patterns of response, although they often differed in the degree of response to light quality. The main response could be related solely to the red: far-red ratio and the calculated phytochrome photoequilibrium. This is the first report of phytochrome-controlled photomorphogenesis in older specimens of a woody perennial. Recommendations for artificial light sources for growing P. radiata and some ecological implications of the results are presented.     

Tree Suppression and the Self-thinning Rule in a Monoculture of Pinus radiata D. Don

An experiment comparing different initial planting densitiesof Pinus radiata D. Don was established in 1940 at Mt. Burrin south-east South Australia. It had been measured periodicallyto age 40 years. It is shown that after some years the treesof each individual plot of the experiment segregated into agroup of ‘suppressed’ trees which grew little indiameter and a group of actively growing ‘dominant’trees. This is consistent with other work that has suggestedthat monocultures tend to produce a two-tiered canopy. The slopeof the relationship between the logarithm of the mean weightper tree and the logarithm of the density of the various plotsof the experiment (the ‘competition-density effect’)was computed at each age at which the experiment was measured,both for all live trees of the plots and for the dominants only.For this analysis tree weights were estimated using the allometricrelationship between weight and diameter at breast height. Forthe dominants, the slope rapidly declined with age until itapproached – 1.5, which is the limiting slope of the competition-densityeffect or the so-called ‘self-thinning rule’. Whenall live trees of the plots were used, the slope declined slowlywith age but remained far from – 1.5. Previously, theself-thinning rule has been seen as determining the way in whichmortality, i.e. self-thinning, occurs in stands. In the presentwork, the rule is found to apply to the larger, actively growingtrees of the stands soon after they start to suppress the smallertrees and long before substantial mortality has occurred inmany stands. Thus tree suppression, rather than mortality, isseen to determine the rule. The pattern of diameter growth oftrees of the various stands of the experiment is found to beconsistent with previous findings that skewness of the frequencydistribution of diameters of trees of stands increases withtime. Pinus radiata D. Don, radiata pine, self-thinning, competition     

The Period of Time Taken by Cambial Derivatives to Grow and Differentiate into Tracheids in Pinus radiata: D. Don

Cambial activity and tracheid development were studied in astand of Pinus radiata D. Don growing in Victoria, Australia.Samples were taken at weekly intervals from 16 August 1966 to19 September 1967. Cell counts were made from these samplesand the results used to estimate the time periods required tocomplete various phases of tracheid development. The average time required for cells of the cambial zone to completea division cycle was about 4 weeks, with little evidence ofany variation during the growing season. The time required forradial growth was about 3 weeks early in the season, graduallyreducing to about 1.5 weeks before the period of dormancy setin. The time required for the deposition of the secondary cellwall was 3 to 4 weeks early in the season, increasing to 8 to10 weeks, later. This increased period was accompanied by areduced rate of deposition of cell-wall material.     

Photosynthesis and wood structure in Pinus radiata D. Don during dehydration and immediately after rewatering

Abstract. Experiments were carried out on Pinus radiata (D. Don) trees grown as cuttings from clonal parent stock. Some of these trees were about 0.4 m high while others were about 5 m high; all were grown in containers. The stem diameters at the tops and at the bottoms of the large trees, rates of photosynthesis, and needle water potentials were measured both when the trees were well watered and as they dehydrated after water was withheld. The water potentials of well-watered plants was highest in the small trees and lowest at the top of the large trees. When water was withheld, photosynthesis was in most cases unaffected by a small reduction in water potential, but the rate of photosynthesis fell as water potentials declined further. The stems of the large trees expanded at a constant rate when the trees were well watered and for part of the dehydration period, while subsequent stem shrinkage and the fall in photosynthesis both occurred at approximately the same time.
Water potentials increased little in the 24 h after rewatering, and significant rates of photosynthesis were not measured until 2 or 3 d later while renewed stem expansion was not measured until 2 d after rewatering.
Water deficits reduced the lumen diameter of newly matured stem tracheids, but increased the thickness of their walls. After 1 month of water potentials of about −2.4 MPa, tracheid lumen diameter and wall thickness were both much reduced, and this reduction continued in tracheids maturing shortly after rewatering.     

The response of photosynthetic model parameters to temperature and nitrogen concentration in Pinus radiata D. Don

Responses of photosynthesis (A) to intercellular CO₂ concentration (c_i) in 2-year-old Pinus radiata D. Don seedlings were measured at a range of temperatures in order to parametrize a biophysical model of leaf photosynthesis. Increasing leaf temperature from 8 to 30°C caused a 4-fold increase in V_cmax, the maximum rate of carboxylation (10.7–43.3 μol m^?2 s^?1 and a 3-fold increase in J_max, the maximum electron transport rate (20.5–60.2 μmol m ^?2 s^?1). The temperature optimum for J_max was lower than that for V_cmax, causing a decline in the ratio J_max:V_cmax from 2.0 to 1.4 as leaf temperature increased from 8 to 30°C. To determine the response of photosynthesis to leaf nitrogen concentration, additional measurements were made on seedlings grown under four nitrogen treatments. Foliar N concentrations varied between 0.36 and 1.27 mol kg^?1, and there were linear relationships between N concentration and both V_cmax and J_max. Measurements made throughout the crown of a plantation forest tree, where foliar N concentrations varied from 0.83 mol kg^?1 near the base to 1.54 mol kg^?1 near the leader, yielded similar relationships. These results will be useful in scaling carbon assimilation models from leaves to canopies.     

Inter- and intraspecific polymorphism at chloroplast SSR loci and the inheritance of plastids in Pinus radiata D. Don

DNA sequence analysis of chloroplast genomes has revealed many short nucleotide repeats analogous to nuclear microsatellites, or simple sequence repeats (SSRs). We designed PCR primers flanking five of these regions identified in the chloroplast sequence from Pinus thunbergii and tested them for amplification in Pinus radiata, P. elliotii, P. taeda, P. strobus, Pseudotsuga menziesii, Cupressus macrocarpa, four New Zealand native conifer species (Podocarpus totara, Podocarpus hallii, Podocarpus nivalis, Agathis australis), and four angiosperms (Vitex lucens, Nestegis cunninghamii, Actinidia chinensis, and Arabidopsis thaliana). A PCR product in the expected size range was amplified from all species and interspecific polymorphism was detected at all five loci. Intraspecific polymorphism was detected in P. radiata with four of the five primer pairs. One of these polymorphic chloroplast SSR (cpSSR) was then used to determine the inheritance of chloroplasts in 206 progeny from four control-pollinated, full-sibling P. radiata families. Approximately 99% of the progeny had the cpSSR variant of the pollen parent indicating that in Pinus radiata, like most other conifers, chloroplasts are typically inherited from the paternal parent. These results suggest that polymorphic chloroplast SSRs will be a valuable tool for studying chloroplast diversity, cyto-nuclear disequilibrium, and plastid inheritance in a range of species, and for the analysis of gene flow via pollen and paternity in species with paternal transmission of chloroplasts.     

Molecular characterization of Nicaraguan Pinus tecunumanii Schw. ex Eguiluz et Perry populations for in situ conservation

Key message

We conducted molecular characterization of Nicaraguan Pinus tecunumanii populations using microsatellite markers. Populations possess considerable genetic variation but there are risks associated with inbreeding and population fragmentation.

Abstract

We carried out a molecular characterization of three natural populations of Pinus tecunumanii using nine microsatellite markers. All studied populations occur in Nicaragua, where the species has declined primarily due to human-influenced factors. The results showed that there is a high amount of genetic variation in populations (expected heterozygosities 0.775–0.841), populations do not show significant differentiation (mean F _ST 0.0073), apparently due to frequent gene flow or a more continuous distribution and homogenous genetic composition in the past, and inbreeding is common in all populations (F _IS 0.705–0.780). The Structure analysis revealed that there is no evident clustering pattern among P. tecunumanii individuals. Although all studied populations possess a considerable amount of genetic variation, risks associated with inbreeding and population fragmentation should be acknowledged and a conservation strategy developed to safeguard the genetic resources of P. tecunumanii.     

A set of microsatellite markers for fingerprinting and breeding applications in Pinus radiata.

Fifty microsatellite markers were developed and characterized in Pinus radiata, and from among these, a subset of 10 easily scored and highly polymorphic markers was selected for use in fingerprinting, quality control, and breeding applications. The markers were characterized based on reliable and reproducible amplification, observed and expected heterozygosities, number of alleles, a low frequency of null alleles, and a lack of close linkage with other selected markers. Allele numbers and frequencies were estimated using 24 first-generation breeding clones from Australia and New Zealand. Observed heterozygosities for the selected markers were all greater than 0.67, and there was an average of 10.5 alleles/locus. The occurrence of null alleles was checked with megagametophytes from mother trees for loci that appeared to be homozygous. The 10 markers are not closely linked (r < 0.20 and LOD > 3) to each other. The selected microsatellites fall into three discrete size classes, and with appropriate selection of fluorescent dyes for 5' end labeling, can be multiplexed with up to 6 markers/sample on an ABI PRISM 310 or similar instrument.     

Response of transpiration and photosynthesis to a transient change in illuminated foliage area for a Pinus radiata D. Don tree

Sudden but transient changes in the fraction or illuminated foliage area in a well-watered 7-year-old Pinus radiata D. Don tree were imposed by completely covering either the upper 22% or the lower 78% of the foliage for periods of up to 36 h. Measurements of transpiration flux density (E), tree conductance (g_t), stomatal conductance (g_s) and net photosynthesis (A) were made to test the hypothesis that compensatory responses would occur in the remaining illuminated foliage when the cover was installed. When the lower foliage was covered there was an immediate decrease in g_t. However, when tree conductance was normalized with respect to the illuminated leaf area (g_t'), it increased between 50 and 75%, depending on the value of air saturation deficit (D). The effect was also apparent from concurrent measurements of increases in g_s and A up to 59 and 24%, respectively, for needles in the top third of (he crown. When the cover was removed these effects were reversed. The changes in the lower foliage when the upper foliage was covered were much smaller. Changes in bulk needle water potential were small. It is suggested that the observed responses occurred because of a perturbation to the hydraulic pathway in the xylem that could have triggered the action of a chemical signal to regulate stomatal conductance and photosynthesis.     

Root Regeneration and Plant Water Status of Pinus radiata D. Don Seedlings Transplanted to Different Soil Temperatures

The regeneration and configuration of new roots on transplanted8 month old Pinus radiata D. Don seedlings were measured inrelation to soil temperature and time after transplanting. Theeffects of root wrenching and nutrition on root regenerationwere compared. Low soil temperature adversely affected the initiationand elongation of new roots. The branching and morphology ofthe new roots were also influenced by soil temperature. Theprocesses of initiation and elongation are greatly retardedbelow a critical root temperature between 11 and 14 ?C. Wrenchingand fertilization stimulated new root growth at low temperatures.There is a close interrelationship between the constituent partsof the network which forms the new root system. The first-orderlaterals on transplants are critical as a basic framework forextension and production of second-order laterals which formthe major part of the regenerating root system. Needle water potential was closely related to the new root growth.The water stress experienced by the transplants for severalweeks after planting was due primarily to the suppressive effectof low soil temperature on root regeneration. Wrenched plantswere less water-stressed than unwrenched plants. Field measurements confirmed the finding from controlled experimentsthat, in southern Australia, the low soil temperature in theplanting season imposes a major restriction on early root regenerationwhich in turn inflicts water stress in transplants. This needsto be considered during the planning of planting and fertilization.