Stem moisture content prediction model for Larix olgensis based on beta regression.

To investigate the longitudinal variation patterns of sapwood, heartwood, bark and stem moisture content along the trunk of artificial Larix olgensis, we constructed mixed effect models of moisture content based on beta regression by combining the effects of sampling plot and sample trees. We used two sampling schemes to calibrate the model, without limiting the relative height (Scheme Ⅰ) and with a limiting height of less than 2 m (Scheme II). The results showed that sapwood and stem moisture content increased gradually along the trunk, heartwood moisture content decreased slightly and then increased along the trunk, and bark moisture content increased along the trunk and then levelled off before increasing. Relative height, height to crown base, stand area at breast height per hectare, age, and stand dominant height were main factors driving moisture content of L. olgensis. Scheme Ⅰ showed the stable prediction accuracy when randomly sampling moisture content measurements from 2-3 discs to calibrate the model, with the mean absolute percentage error (MAPE) of up to 7.2% for stem moisture content (randomly selected 2 discs), and the MAPE of up to 7.4%, 10.5% and 10.5% for sapwood, heartwood and bark moisture content (randomly selected 3 discs), respectively. Scheme Ⅱ was appropriate when sampling moisture content measurements from discs of 1.3 and 2 m height and the MAPE of sapwood, heartwood, bark and stem moisture content reached 7.8%, 11.0%, 10.4% and 7.1%, respectively. The prediction accuracies of all mixed effect beta regression models were better than the base model. The two-level mixed effect beta regression models, considering both plot effect and tree effect, would be suitable for predicting moisture content of each part of L. olgensis well.

Carbon stock projection for four major forest plantation species in Japan.

Carbon sequestration via afforestation and forest growth is effective for mitigating global warming. Accurate and robust information on forest growth characteristics by tree species, region, and large-scale land-use change is vital and future prediction of forest carbon stocks based on this information is of great significance. These predictions allow exploring forestry practices that maximize carbon sequestration by forests, including wood production. Forest inventories based on field measurements are considered the most accurate method for estimating forest carbon stocks. Japan's national forest inventories (NFIs) provide stand volumes for all Japanese forests, and estimates from direct field observations (m-NFIs) are the most reliable. Therefore, using the m-NFI from 2009 to 2013, we selected four major forest plantation species in Japan: Cryptomeria japonica, Chamaecyparis obtusa, Pinus spp., and Larix kaempferi and presented their forest age-carbon density function. We then estimated changes in forest carbon stocks from the past to the present using the functions. Next, we investigated the differences in the carbon sequestration potential of forests, including wood production, between five forestry practice scenarios with varying harvesting and afforestation rates, until 2061. Our results indicate that, for all four forest types, the estimates of growth rates and past forest carbon stocks in this study were higher than those considered until now. The predicted carbon sequestration from 2011 to 2061, assuming that 100 % of harvested carbon is retained for a long time, twice the rate of harvesting compared to the current rate, and a 100 % afforestation rate in harvested area, was three to four times higher than that in a scenario with no harvesting or replanting. Our results suggest that planted Japanese forests can exhibit a high carbon sequestration potential under the premise of active management, harvesting, afforestation, and prolonging the residence time of stored carbon in wood products with technology development.

Boreal conifers maintain carbon uptake with warming despite failure to track optimal temperatures.

Warming shifts the thermal optimum of net photosynthesis (ToptA) to higher temperatures. However, our knowledge of this shift is mainly derived from seedlings grown in greenhouses under ambient atmospheric carbon dioxide (CO2) conditions. It is unclear whether shifts in ToptA of field-grown trees will keep pace with the temperatures predicted for the 21st century under elevated atmospheric CO2 concentrations. Here, using a whole-ecosystem warming controlled experiment under either ambient or elevated CO2 levels, we show that ToptA of mature boreal conifers increased with warming. However, shifts in ToptA did not keep pace with warming as ToptA only increased by 0.26-0.35 °C per 1 °C of warming. Net photosynthetic rates estimated at the mean growth temperature increased with warming in elevated CO2 spruce, while remaining constant in ambient CO2 spruce and in both ambient CO2 and elevated CO2 tamarack with warming. Although shifts in ToptA of these two species are insufficient to keep pace with warming, these boreal conifers can thermally acclimate photosynthesis to maintain carbon uptake in future air temperatures.

The shifts in the structure of the prokaryotic community of mountain-grassland soil under the influence of artificial larch plantations.

Creation of artificial forest plantations on a global scale is one of the ways to mitigate the negative effects of climate change on ecosystems, at the same time providing soil protection from erosion, regulation of the hydrological regime and carbon sequestration in soils of different natural and climatic zones. However, the change of the dominant plant community cause significant ecosystem changes, reflecting at the structure and functioning of the soil microbial complex as well. The shifts in prokaryotic community of the meadow soil resulting from the conversion of the native meadow (further grassland) phytocenosis to the artificial forest plantations was investigated with the use of NGS sequencing technology and metabarcoding approach-amplicon sequencing of V4 region of 16 S rRNA (performed on Illumina Miseq platform). The identified shifts in taxonomic structure and diversity may be the result of changes in the physic-chemical conditions of soils and, on the other hand, may serve as indicators of such changes. Cultivation of larch led to an increase in the diversity of the prokaryotic community and its stratification by depth. The acidifying effect of larch manifested itself in an increase in the proportion and diversity of acidobacteria, in the abundance of oligotrophic microorganisms of phyla Chloroflexi, Firmicutes, and a simultaneous comparative decrease in the bacteria of Verrucomicrobia phylum, alphaproteobacteria of or. Rhizobiales and Burkholderiales. The absence of clearly expressed dominants in the prokaryotic community, as well as a significant increase in alpha-diversity indices, compared with the control plot of native mountain-meadow soil under grassland vegetation, suggests a transitional nature of the soil ecosystem of artificial forest plantations.

Embryogenic callus induction from immature zygotic embryos and genetic transformation of Larix kaempferi 3x Larix gmelinii 9.

To date, there are few reports of the successful genetic transformation of larch and other conifers, mainly because it is difficult to transform and integrate exogenous genes. In this study, hybrid larch Larix kaempferi 3x Larix gmelinii 9 cones were collected on June 27, July 1, July 4, July 7 and July 16, 2017. Embryogenic callus induction was studied using a combination of different plant growth regulators and concentrations. The results showed that July 1 was the best stage; the highest induction rate was 10.83%, which cultured in BM medium (Button medium, which formula was listed in S1 Table) with 1.0 mg/L 2,4-D (2,4-dichlorophenoxyacetic acid) and 0.2 mg/L KT(kinetin). When cultured on a proliferation medium for 12 days, proliferation was the fastest, reaching 323.08%, which could also maintain the freshness and vitality. The suitable pre-culture medium for somatic embryogenesis was 1/4 BM medium containing 10 g/L inositol and 60 g/L sucrose. The combination of 45 mg/L ABA (abscisic acid) and 75 g/L PEG4000 (Polyethyene glycol 4000) could promote the number of somatic embryos, and reached the maximum, 210 140 per 1 g FW. The genetic transformation was carried out by the Agrobacterium-mediated transformation method with embryogenic callus cultured for 12 days. The results showed the optimal OD600 of the infection solution(suspension of A. tumefaciens) was 0.5, co-culture time was 2 days, and screening concentration of Hyg (hygromycin B) was 4 mg/L. In this study, the transformation rate of resistance callus was 32.1%. It provides a reference for low genetic transformation efficiency of larch at present. This study could be beneficial for the innovation and breeding of larch by genetic engineering and provides a certain basis for rapid propagation of excellent larch germplasm resources and genetic engineering breeding of larch and other conifers.

Root foraging of birch and larch in heterogeneous soil nutrient patches under water deficit.

Water and nutrient are two critical factors that limit plant growth to spatial-temporal extents. Tree root foraging behavior has not received adequate attention in heterogeneous soil environments in temperate forest under drought pressure. In this study, birch (Betula platyphylla) and larch (Larix olgensis) seedlings were raised in pots in a split-root system with artificially heterogeneous soil environments to study the root foraging response to drought. Potted space was split into two halves where substrates were mixed with fertilizers in 67.5 mg nitrogen (N) plant-1 (N-P2O5-K2O, 14-13-13) to both halves as to create a homogeneous condition. Otherwise, a rate of 135 mg N plant-1 of fertilizers was delivered to a random half to create a heterogeneous condition. Half of seedlings were fully sub-irrigated every three days with the other half received the drought treatment by being watered every six days. Both birch and larch seedlings showed greater net shoot growth and biomass increment in well-watered condition, while root morphology was promoted by drought. Both species placed more fine roots with higher root N concentration in nutrient-enriched patches. In the heterogeneous pattern, birch showed a higher foraging precision assessed by biomass and greater foraging plasticity assessed in morphology and physiology. In contrast, larch seedlings had higher root N concentration in the well-watered condition. Neither species showed a significant response of N utilization to the heterogeneous pattern, but both used more N when water supply was improved. Overall, birch is better at acclimating to heterogeneous soil conditions, but its ability to seize N was lower than larch when drought was alleviated.

Spatial and Temporal Distribution of Arabinogalactan Proteins during Larix decidua Mill. Male Gametophyte and Ovule Interaction.

The role of ArabinoGalactan Proteins (AGPs) in the sexual reproduction of gymnosperms is not as well documented as that of angiosperms. In earlier studies, we demonstrated that AGPs play important roles during ovule differentiation in Larix decidua Mill. The presented results encouraged us to carry out further studies focused on the functions of these unique glycoproteins during pollen/pollen tube and ovule interactions in Larix. We identified and analyzed the localization of AGPs epitopes by JIM4, JIM8, JIM13 and LM2 antibodies (Abs) in male gametophytes and ovule tissue during pollination, the progamic phase, and after fertilization and in vitro growing pollen tubes. Our results indicated that (1) AGPs recognized by JIM4 Abs play an essential role in the interaction of male gametophytes and ovules because their appearance in ovule cells is induced by physical contact between reproductive partners; (2) after pollination, AGPs are secreted from the pollen cytoplasm into the pollen wall and contact the extracellular matrix of stigmatic tip cells followed by micropylar canal cells; (3) AGPs synthesized in nucellus cells before pollen grain germination are secreted during pollen tube growth into the extracellular matrix, where they can directly interact with male gametophytes; (4) in vitro cultured pollen tube AGPs labeled with LM2 Abs participate in the germination of pollen grain, while AGPs recognized by JIM8 Abs are essential for pollen tube tip growth.

Cavitation fatigue in conifers: a study on eight European species.

After drought-induced embolism and repair, tree xylem may be weakened against future drought events (cavitation fatigue). As there are few data on cavitation fatigue in conifers available, we quantified vulnerability curves (VCs) after embolism/repair cycles on eight European conifer species. We induced 50% and 100% loss of conductivity (LC) with a cavitron, and analyzed VCs. Embolism repair was obtained by vacuum infiltration. All species demonstrated complete embolism repair and a lack of any cavitation fatigue after 50% LC . After 100% LC, European larch (Larix decidua), stone pine (Pinus cembra), Norway spruce (Picea abies), and silver fir (Abies alba) remained unaffected, while mountain pine (Pinus mugo), yew (Taxus baccata), and common juniper (Juniperus communis) exhibited 0.4-0.9 MPa higher vulnerability to embolism. A small cavitation fatigue observed in Scots pine (Pinus sylvestris) was probably biased by incomplete embolism repair, as indicated by a correlation of vulnerability shifts and conductivity restoration. Our data demonstrate that cavitation fatigue in conifers is species-specific and depends on the intensity of preceding LC. The lack of fatigue effects after moderate LC, and relevant effects in only three species after high LC, indicate that conifers are relatively resistant against cavitation fatigue. This is remarkable considering the complex and delicate conifer pit architecture and may be important considering climate change projections.

Long-term effects of mixed planting on arbuscular mycorrhizal fungal communities in the roots and soils of Juglans mandshurica plantations.

BACKGROUND: Establishing mixed plantations is an effective way to improve soil fertility and increase forest productivity. Arbuscular mycorrhizal (AM) fungi are obligate symbiotic fungi that can promote mineral nutrient absorption and regulate intraspecific and interspecific competition in plants. However, the effects of mixed plantations on the community structure and abundance of AM fungi are still unclear. Illumina MiSeq sequencing was used to investigate the AM fungal community in the roots and soils of pure and mixed plantations (Juglans mandshurica × Larix gmelinii). The objective of this study is to compare the differential responses of the root and rhizosphere soil AM fungal communities of Juglans mandshurica to long-term mixed plantation management. RESULTS: Glomus and Paraglomus were the dominant genera in the root samples, accounting for more than 80% of the sequences. Compared with that in the pure plantation, the relative abundance of Glomus was higher in the mixed plantation. Glomus, Diversispora and Paraglomus accounted for more than 85% of the sequences in the soil samples. The relative abundances of Diversispora and an unidentified genus of Glomeromycetes were higher and lower in the pure plantation, respectively. The Root_P samples (the roots in the pure plantation) had the highest number of unique OTUs (operational taxonomic units), which belonged mainly to an unidentified genus of Glomeromycetes, Paraglomus, Glomus and Acaulospora. The number of unique OTUs detected in the soil was lower than that in the roots. In both the root and soil samples, the forest type did not have a significant effect on AM fungal diversity, but the Sobs value and the Shannon, Chao1 and Ace indices of AM fungi in the roots were significantly higher than those in the soil. CONCLUSIONS: Mixed forest management had little effect on the AM fungal community of Juglans mandshurica roots and significantly changed the community composition of the soil AM fungi, but not the diversity.

Transcription factor LkWOX4 is involved in adventitious root development in Larix kaempferi.

WUSCHEL-related homeobox4 (WOX4) plays important roles in vascular formation and adventitious root (AR) development. Here, we cloned the WOX4 from the AR of Larix kaempferi, whose cDNA is 1452 bp in length and encodes 483 amino acids. LkWOX4 is mainly expressed in the layer formation area of the stem at 10 days after cutting and its expression levels in the middles and ends of the ARs were higher than that in the AR tips. The fused protein LkWOX4-GFP localized in the nucleus. The heterologous overexpression of LkWOX4 in 84 K poplar significantly increased AR numbers and decreased AR lengths. In LkWOX4 plants, the endogenous jasmonic acid and abscisic acid contents significantly decreased in stems, while the auxin, jasmonic acid and abscisic acid contents significantly increased in ARs. RNA-Seq of those LkWOX4 overexpression poplar plants showed that the expression of plant hormone signaling genes (ARF2, ARF3, ARF7 and ARF18), rooting-related transcription factors (WOX5, LBD29 and SCR) and root development-related genes (CYCD3, GRF1 and TAA1) were affected. Moreover, we found that LkWOX4 interacts with LkPAT18, LkACBP6, and LkCIP7 using yeast two hybrid screening. Thus, we found LkWOX4 involves in the AR initiation and development, which might be regulated through the IAA, JA and ABA signaling pathways.

Nitrogen loading increases the ozone sensitivity of larch seedlings with higher sensitivity to nitrogen loading.

Larch (Larix sp.) tree is a critical species for the future afforestation in Northeast Asia. The impacts of elevated concentrations of ground-level ozone (O3) and nitrogen (N) deposition are raising concerns. However, knowledge of the combined effects of elevated O3 and N loading are still limited. We investigated whether nitrogen loading mitigates the negative impacts of ozone on two larch species: the Japanese larch (L. kaempferi) and its hybrid larch F1 (L. gmelinii var. japonica × L. kaempferi) or not. We used open-top cambers and compared responses of the larch seedlings. Results showed the N loading mitigated the negative effects of O3 on Japanese larch. However, in hybrid larch F1, N loading did not mitigate O3-induced inhibition of growth and photosynthetic capacity. Mitigation effect of N loading on negative O3 impacts may vary between the two Larix spp., Hybrid larch F1 could be more affected by the combined effects of O3 and N loading due to its higher growth response to N loading. Elevated O3 also reduced leaf nitrogen/phosphorus (N/P) ratio by elevated O3, with significant effects in hybrid larch F1, particularly under N loading. In the present study, leaf N/P ratio was utilized to validate the hypothesis that a positive effect of N loading may be observed if O3 does not induce P limitation in Larix spp. We demonstrated a potential leaf N/P ratio function, which could reflect responses to O3 and N loading in hybrid larch F1.

Isolation and characterization of larch BABY BOOM2 and its regulation of adventitious root development.

The BABY BOOM2 gene, designated LkBBM2, and its promoter were isolated from hybrid larch (Larix kaempferi × L. olgensis). The open reading frame of LkBBM2 was 2574 bp, encoding 857 amino acids. The LkBBM2 protein contains two AP2 DNA binding domains and a BBM specific motif, but lacks the euANT5 motif common to AP2 family members. The LkBBM2 promoter contains several hormone response and root-specific expression elements. LkBBM2 expression was significantly higher in larch adventitious roots (ARs) than in stems, leaves or stem tips, and increased after auxin treatment. The fused protein LkBBM2-GFP was localized in both the nucleus and cytoplasm whereas LkBBM1-GFP was only localized in the nucleus. Over-expression of LkBBM2 and LkBBM1 in Arabidopsis significantly elongated the roots. Furthermore, over-expression those two genes in the hybrid poplar (Populus alba × P. glandulosa) significantly increased ARs number. We speculated that these two genes regulate AR development.

Couplings in cell differentiation kinetics mitigate air temperature influence on conifer wood anatomy.

Conifer trees possess a typical anatomical tree-ring structure characterized by a transition from large and thin-walled earlywood tracheids to narrow and thick-walled latewood tracheids. However, little is known on how this characteristic structure is maintained across contrasting environmental conditions, due to its crucial role to ensure sap ascent and mechanical support. In this study, we monitored weekly wood cell formation for up to 7 years in two temperate conifer species (i.e., Picea abies (L.) Karst and Larix decidua Mill.) across an 8°C thermal gradient from 800 to 2,200 m a.s.l. in central Europe to investigate the impact of air temperature on rate and duration of wood cell formation. Results indicated that towards colder sites, forming tracheids compensate a decreased rate of differentiation (cell enlarging and wall thickening) by an extended duration, except for the last cells of the latewood in the wall-thickening phase. This compensation allows conifer trees to mitigate the influence of air temperature on the final tree-ring structure, with important implications for the functioning and resilience of the xylem to varying environmental conditions. The disappearing compensation in the thickening latewood cells might also explain the higher climatic sensitivity usually found in maximum latewood density.

Microbial biomass and enzymatic responses to temperate oak and larch forest thinning: Influential factors for the site-specific changes.

Microbial biomass and enzyme activity are essential for ecosystem function in managed forests; however, uncertainty remains because microbial biomass and enzymatic responses to thinning highly differ with case studies. This study addressed the drivers for the site-specific responses of microbial biomass and enzyme activity to thinning. Study sites included two oak and three larch forests; each had un-thinned control, intermediate thinning (15-23% basal area reduction), and heavy thinning treatments (30-44% basal area reduction). Soil properties (temperature, water content, pH, total and inorganic nitrogen, and total carbon/nitrogen ratio), microbial biomass, enzyme (ß-glucosidase, N-acetylglucosaminidase, leucyl aminopeptidase, acid phosphatase, and phenol oxidase) activity, and soil carbon storage were determined 6 years after thinning. Compared to the control, microbial biomass carbon and nitrogen were higher under the intermediate and the heavy thinning by 13.9 and 24.4% and 11.5 and 29.9% at one oak forests, respectively, and higher under the intermediate thinning by 53.7 and 70.7% at one larch forests. There were the post-thinning changes in leucyl aminopeptidase activity by -46.9% and by 150.0-210.0% at an oak and larch forest, respectively, acid phosphatase activity by 60.0% at one oak forest, and phenol oxidase activity by 355.0% at one oak forest. The effect sizes of thinning for soil properties explained 94% and 77% of variance of the effect sizes for microbial biomass and enzyme activity. Especially, the effect sizes for soil water content, NH4+, total carbon/nitrogen ratio, and temperature were the most influential. Furthermore, the effect size for soil carbon storage was parabolically related to the effect size for microbial biomass carbon (R2 = 0.66). These findings highlight that inconsistent thinning effects on soil properties varied microbial biomass and enzymatic responses to thinning, which differentiated the change in soil carbon storage across sites. Future studies should consider such inconsistencies when examining the effects of forest management.

Deciphering Hybrid Larch Reaction Norms Using Random Regression.

The link between phenotypic plasticity and heterosis is a broad fundamental question, with stakes in breeding. We report a case-study evaluating temporal series of wood ring traits of hybrid larch (Larix decidua × L. kaempferi and reciprocal) in relation to soil water availability. Growth rings record the tree plastic responses to past environmental conditions, and we used random regressions to estimate the reaction norms of ring width and wood density with respect to water availability. We investigated the role of phenotypic plasticity on the construction of hybrid larch heterosis and on the expression of its quantitative genetic parameters. The data came from an intra-/interspecific diallel mating design between both parental species. Progenies were grown in two environmentally contrasted sites, in France. Ring width plasticity with respect to water availability was confirmed, as all three taxa produced narrower rings under the lowest water availability. Hybrid larch appeared to be the most plastic taxon as its superiority over its parental species increased with increasing water availability. Despite the low heritabilities of the investigated traits, we found that the expression of a reliable negative correlation between them was conditional to the water availability environment. Finally, by means of a complementary simulation, we demonstrated that random regression can be applied to model the reaction norms of non-repeated records of phenotypic plasticity bound by a family structure. Random regression is a powerful tool for the modeling of reaction norms in various contexts, especially perennial species.

[Change pattern of heartwood of Larix olgensis plantation.] / 长白落叶松人工林心材变化规律.

Based on the data of 1179 discs and whorls of 49 trees from larch (Larix olgensis) plantations located in Mengjiagang forest farm in Heilongjiang Province, China, we analyzed the longitudinal variation pattern of heartwood radius. The results showed that the heartwood radius decreased with the increases of tree height, which was basically the same as the trunk shape. The relationship between the xylem radius (XR), diameter at breast height (DBH) and cambial age (CA) with the heartwood radius was significant. The stepwise regression analysis was used to develop heartwood radius (HR) and heartwood area (HA) models: HR=b1+b2XR2+b3CA+b4XR, HA=b1+b2DBH·XR+b3CA+b4DBH·XR2. We used the evaluation statistics such as AIC, BIC, Log Likelihood and Likelihood ratio test to compare the heartwood radius and heartwood area models which fitted with the plot effect and tree effect. The heartwood radius and heartwood area models with parameters b1, b2, b3 as mixed effects performed best when the tree effect was considered. The prediction accuracy of the mixed model was better than that of the basic model. In the application, the total heartwood radius and area could be predicted by the mixed model. Beta regression model was used to simulate the heartwood proportion. In this model, all parameters were significant, and the coefficients of determination were relatively high, with a good simulation effect.

[Reconstruction of July NDVI over 172 years based on tree-ring width of Larix chinensis in Taibai Mountain Nature Reserve, China.] / åŸºäºŽå¤ªç™½çº¢æ‰æ ‘è½®å®½åº¦é‡å»ºè¿‘172年太白山自然保护区7月NDVI.

We used the correlation analysis of Larix chinensis tree-ring width chronologies, meteorological data as well as regional NDVI data to reconstruct the NDVI sequence variation of the north and south aspects of Taibai Mountain Nature Reserve in the recent 172 years. The results showed that the NDVI variation of Taibai Mountain Nature Reserve was consistent with plant growth dyna-mics. Hydrothermal condition in growing season was the dominant factor controlling NDVI changes. Tree-ring width was significantly positively correlated with NDVI in growing season. The strongest correlation occurred in July. Thus, we used the long time series of tree ring width index in July to reconstruct the regional historical period variation of NDVI sequence. The results demonstrated that historical changes of July NDVI sequence existed five dense periods and five sparse periods. Further analysis indicated that these periods corresponded with the climate change trends and drought events. The reconstructed July NDVI sequence of both northern and southern slopes of Taibai Mountain Nature Reserve highlighted the existence of quasi periodic variation of 60 years.

Water stress limits transpiration and growth of European larch up to the lower subalpine belt in an inner-alpine dry valley.

Climate change will further constrain water availability in dry inner-alpine environments and affect water relations and growth conditions in mountain forests, including the widespread larch forests. To estimate the effects of climate conditions on water balance and growth, variation in sap flow and stem radius of European larch was measured for 3 yr along an elevation transect from 1070 to 2250 m above sea level (asl) in an inner-alpine dry valley in South Tyrol/Italy. Additionally, long-term climate-growth relations were derived from tree cores. Sap flow and radial growth were reduced in dry periods up to an elevation of 1715 m, leading to maximum annual growth at 2000 m. In a wet year no growth difference between elevations was observed. Long-term tree ring data showed a positive growth response to precipitation up to 1715 m and to temperature only above 2000 m. Our results demonstrate that reduced water availability and higher atmospheric water demand limit larch at low elevation within dry Alpine regions. This indicates a general upward shift of this species' elevational amplitude upon climate change, and respective negative effects on future silvicultural use and ecosystem services at lower elevations in the European Alps.

LaMIR166a-mediated auxin biosynthesis and signalling affect somatic embryogenesis in Larix leptolepis.

Somatic embryogenesis (SE) involves complex molecular signalling pathways. Understanding molecular mechanism of SE in Larix leptolepis (L. leptolepis) can aid research on genetic improvement of gymnosperms. Previously, we obtained five LaMIR166a (miR166a precursor) -overexpression embryonic cell lines in the gymnosperm Larix leptolepis. The proliferation rates of pro-embryogenic masses in transgenic and wild-type lines were calculated. Overexpression of the miR166a precursor LaMIR166a led to slower proliferation. When pro-embryogenic masses were transferred to maturation medium, the relative expression of LaMIR166a and miR166a in the LaMIR166a-overexpression lines was higher than in the wild-type during SE, while LaHDZ31-34 expression levels also increased without negative control by miR166, suggesting that regulation of HD-ZIP III by miR166a exits stage-specific characteristics. The key indole-3-acetic acid (IAA) biosynthetic gene Nitrilase of L. leptolepis (LaNIT) was identified and the effects of miR166a on auxin biosynthesis and signalling genes were studied. During SE, LaNIT, Auxin response factor1 (LaARF1) and LaARF2 mRNA levels and IAA contents were markedly higher in LaMIR166a-overexpression lines, which revealed lower deformity rate of embryos, indicating endogenous IAA synthesis is required for somatic embryo maturation in L. leptolepis. Additionally, the IAA biosynthesis and signalling genes showed similar expression patterns to LaHDZ31-34, suggesting HD-ZIP III genes have a positive regulatory effect on LaNIT. Our results suggest miR166a and LaHDZ31-34 have important roles in auxin biosynthesis and signalling during SE, which might determine if the somatic embryo normally developed to mature in L. leptolepis.