Roots are vital organs for resource uptake. However, the knowledge regarding the extent by which responses in roots influence plant resistance is still poorly known. In this study, we examined the morphological and physiological responses of lateral roots of Taxodium hybrid ‘Zhongshanshan 406’ (Taxodium mucronatum♀ and Taxodium distichum♂, T. 406) to 8 (DS-8) and 12 days (DS-12) drought. Control plants (CK-8 and CK-12) were well-watered throughout the experiment. Results indicated that drought resulted in significantly decreased root length, surface area, volume, and biomass and a relatively high death rate of roots (>2 mm). Specific root length (SRL) and specific root surface area (SRA) of drought-stressed T. 406 plants were reduced to enhance resource uptake. Meanwhile, root relative water content (RWC) of T. 406 plants in CK-12 treatment was 5.81 times of those in DS-12 treatment. Under drought stress and root superoxide dismutase and ascorbic acid (ASA) activities, proline and hydrogen peroxide (H2O2) contents consistently increased to benefit the elimination of O2−. At the ultrastructural level, the organelle structure of T. 406 plant root tip was visibly damaged because of dehydration. The nucleus swelled and then exhibited uncommon features of disorganization and disruption. In short, our results provided substantial information about lateral root traits of T. 406 plants in response to drought stress, which is crucial to improve the drought resistance of Taxodium hybrid in the future breeding.

As a subfamily of the APETALA 2/ethylene response element binding protein (AP2/EREBP) transcription factor superfamily, the ethylene response factor (ERF) is widely involved in the regulation of growth and response to various abiotic stresses in plants, and has been shown to be the main transcription factor regulating transcription of the genes related to hypoxia and waterlogging stress. In this study, three ThERF genes, with significant differences in expression profile in response to flooding stress, were identified from the transcriptomics data acquired from Taxodium hybrid ‘Zhongshanshan 406’ (T. mucronatum Tenore × T. distichum (L.) Rich) under waterlogging stress: ThERF15, ThERF39 and ThRAP2.3 (GenBank ID: KY463467, KY463468 and KY463470, respectively).The full-length cDNA of each of the three ERFs was obtained using the RACE (rapid amplification cDNA ends) method, and all three were intron-free. Multiple protein sequence alignments indicated that ThERF15, ThERF39 and ThRAP2.3 proteins all had only one AP2-ERF domain and belonged to the ERF subfamily. A transient gene expression assay demonstrated that ThERF15, ThERF39 and ThRAP2.3 were all localized to the nucleus. Real-time quantitative PCR (qPCR) revealed that the expression of ThERF15, ThERF39 and ThRAP2.3 exhibited significant differences, compared with the control, in response to two levels of flooding treatment (half-flooding or total-submergence) of ‘Zhongshanshan 406’. Quantification of ethylene concentration revealed that ethylene was more relevant to the level of expression than the period of flooding treatment. Based on the experimental results above, ThERF15, ThERF39 and ThRAP2.3 were identified as being related to the regulation of downstream flooding- responsive gene expression in ‘Zhongshanshan 406’. ThRAP2.3 is most likely to be a key downstream-response ERF gene to respond to the output of the ethylene signal generated by flooding stress.

模拟江苏省沿海地区盐渍土的离子种类及含量,利用土培法对法青Viburnum odoratissimu、海桐Pittosporum tobira和海滨木槿Hibiscus hamabosieb 3种常用绿化灌木进行0.0(CK)、3.0、5.0和8.0 g·kg-1混合盐胁迫处理,研究其叶绿素、Na+、K+及叶片光合系统的响应.结果表明,3种灌木根、茎、叶中Na+含量在受胁迫后都有增长的趋势,且在根中积累的Na+比茎、叶部分更多,其中海滨木槿最为显著.根系对K+的吸收作用在较低盐浓度下(3.0 g·kg-1)即被抑制,而茎、叶对K+的吸收随混合盐浓度的升高也逐步降低.法青、海桐和海滨木槿受盐胁迫后的K+/Na+均为:叶>茎>根,在相同盐胁迫条件下,海滨木槿、海桐的根、茎、叶均具有更高的K+/Na+比.随着混合盐浓度的升高,3种灌木叶绿素含量、净光合速率、气孔导度及蒸腾速率基本呈下降趋势,法青下降更为明显.结果显示,3种灌木均具有一定的耐盐性,其中海滨木槿耐盐性最强,海桐次之.

‘Zhongshanshan’ is the general designation for the superior interspecific hybrid clones of Taxodium species, which is widely grown for economic and ecological purposes in southern China. Growth is the priority objective in ‘Zhongshanshan’ tree improvement. A high-density linkage map is vital to efficiently identify key quantitative trait loci (QTLs) that affect growth.

[目的]探究中山杉407(Taxodium mucronatum♀ ×T.distichum♂)(T.hybrid'Zhongshanshan 407')的ThP5CS和Thδ-OAT基因与植物耐旱性的关系,为落羽杉属植物抗性育种提供新的基因资源.[方法]利用RACE技术从中山杉407中分别克隆ThP5CS和Thδ-OAT基因cDNA全长,通过生物信息学手段分析预测其编码蛋白的结构和功能.基于干旱-复水试验,采用半定量和实时荧光定量技术研究中山杉407 ThP5CS和Thδ-OAT的表达特性.[结果]从中山杉407中克隆了编码P5 CS和δ-OAT蛋白的基因,分别命名为ThP5CS和Thδ-OAT,其中,Thδ-OAT包含1个长度为1494 bp的ORF,编码497个氨基酸,与樟子松(Pinus sylvestris L.)δ-OAT蛋白的序列相似性高达92％;ThP5CS包含1个长度为1545 bp的ORF,编码514个氨基酸,与花烛属植物(Anthurium amnicola)P5CS蛋白的序列相似性为87％.半定量与定量PCR结果一致,在自然干旱和复水环境下,ThP5CS在中山杉407及其父本落羽杉(T.distichum)、母本墨杉(T.mucronatum)中均表现为先上调再下调的趋势,而Thδ-OAT在中山杉407及其父母本中相对表达量则存在差异.[结论]干旱胁迫下,Thδ-OAT和ThP5CS基因的正反方向调节是中山杉407及其亲本在干旱胁迫与恢复状态下控制脯氨酸水平的关键机制,其中,ThP5CS在中山杉407及其亲本脯氨酸的合成过程中起重要作用.

ERF (Ethylene responsive factor)转录因子是植物AP2/EREBP转录因子超家族的一个亚家族,广泛参与植物生长发育及各种逆境胁迫反应的调控,ERF亚家族中的第Ⅶ类成员(ERF-Ⅶs)己被证实是调节低氧相关基因表达和响应水淹胁迫的主要转录因子.本研究从中山杉406 (Taxodium‘ Zhongshanshan 406’)淹水胁迫下获得的转录组数据中,筛选分离出存在显著差异表达的ERF基因,以中山杉406的根为材料,采用RACE技术克隆获得1个ERF-Ⅶ类基因ThRAP2.1 (GenBank登录号为KY463469).生物信息学分析表明,ThRA P2.1全长为1 024 bp,包含735bp的开放阅读框,编码244个氨基酸,无内含子,蛋白质分子量为27.14kD,等电点为9.30.原生质体的瞬时表达显示:ThRAP2.1蛋白定位于细胞核.实时荧光定量PCR显示:ThRA P2.1基因的表达量在中山杉406淹水后产生显著差异,全淹及半淹处理下,ThRA P2.1基因在根中的表达量均显著高于对照,在叶中的表达量均显著低于对照.上述实验结果表明ThRA P2.1参与了中山杉406在水淹胁迫响应中的调控,可作为候选基因用于中山杉及其他落羽杉属树木耐水淹机制的研究.

This work provides a guideline for application of Taxodium ‘Zhongshanshan’ reference gene usage in the qRT-PCR system, and also for reference gene selection for other Taxodiaceae species.

By simulating ion components of coastal saline soil, relative electric conductivity, malondialdehyde ( MDA ) content, and antioxidant enzyme activities in leaves, and absorption and distribution of Na+ and K+ in different organs of three arbor species of Michelia compressa ( Maxim.) Sarg., Taxodium hybrid 'Zhongshanshan 406 ', and Ligustrum lucidum Ait. under different concentrations (3, 5, and 8 g·kg-1) of mixed salt stresses were investigated. The results show that three arbor species in each treatment group all survive except for L. lucidum treated with 8 g·kg-1 of mixed salt stress. Both relative electric conductivity and MDA content in leaves of three arbor species increase with enhancing of mixed salt stress concentration. Relative electric conductivity in leaves of three arbor species in each treatment group from high to low is M. compressa, L. lucidum, T. hybrid'Zhongshanshan 406' , while their MDA content is on the contrary basically. With enhancing of mixed salt concentration, peroxidase ( POD) activity in leaves of M. compressa increases significantly, and its catalase ( CAT) activity in 5 and 8 g · kg-1 mixed salt stress treatment groups are significantly higher than that in 3 g·kg-1 mixed salt stress treatment group and the control group (0 g·kg-1 mixed salt), while there is no significant difference in superoxide dismutase ( SOD) activity among different treatment groups. SOD and CAT activities in leaves of T. hybrid 'Zhongshanshan 406' in 8 g·kg-1 mixed salt stress treatment group are significantly higher than those in 3 and 5 g · kg-1 mixed salt stress treatment groups and the control group;while there is no significant difference in POD activity among 3, 5, and 8 g·kg-1 mixed salt stress treatment groups, but their POD activity is higher than that in the control group. There is no significant change in SOD, POD, and CAT activities in leaves of L. lucidum under mixed salt stress. With enhancing of mixed salt stress concentration, Na+ content in roots, stems, and leaves of three arbor species shows an upward trend in general, K+ content in different organs of three arbor species shows different changing trends, and K+/Na+ ratio shows a downward trend in general. K+/Na+ ratio in roots, stems, and leaves of T. hybrid 'Zhongshanshan 406' in each treatment group is generally higher than that in M. compressa and L. lucidum. In general, T. hybrid 'Zhongshanshan 406 ' has the best salt tolerance among three arbor species tested, indicating that it is the optimal tree species for saline land improvement, and afforestation and greening in coastal areas.

Among the GRAS family of transcription factors, SHORT ROOT (SHR) and SCARECROW (SCR) are key regulators of the formation of root tissues. In this study, we isolated and characterized two genes encoding SHR proteins and one gene encoding an SCR protein: ThSHR1 (Accession Number MF045148), ThSHR2 (Accession Number MF045149) and ThSCR (Accession Number MF045152) in the adventitious roots of Taxodium hybrid 'Zhongshanshan'.Gene structure analysis indicated that ThSHR1, ThSHR2 and ThSCR are all intron free. Multiple protein sequence alignments showed that each of the corresponding proteins, ThSHR1, ThSHR2 and ThSCR, contained five well-conserved domains: leucine heptad repeat I (LHRI), the VHIID motif, leucine heptad repeat II (LHR II), the PFYRE motif, and the SAWmotif. The phylogenetic analysis indicated that ThSCR was positioned in the SCR clade with the SCR proteins from eight other species, while ThSHR1 and ThSHR2 were positioned in the SHR clade with the SHR proteins from six other species. Temporal expression patterns of these genes were profiled during the process of adventitious root development on stem cuttings. Whereas expression of both ThSHR2 and ThSCR increased up to primary root formation before declining, that of ThSHR1 increased steadily throughout adventitious root formation. Subcellular localization studies in transgenic poplar protoplasts revealed that ThSHR1, ThSHR2 and ThSCR were localized in the nucleus. Collectively, these results suggest that the three genes encode Taxodium GRAS family transcription factors, and the findings contribute to improving our understanding of the expression and function of SHR and SCR during adventitious root production, which may then be manipulated to achieve high rates of asexual propagation of valuable tree species.

The physiological acclimation of Taxodium hybrid ‘zhongshanshan 118’ ( T .118) plants to a progressive drought stress and drought-stressed to recovery treatment (DS-R) was investigated in this study. Plants of control (C) treatment were watered daily throughout the experiment. Results indicated that water deficit reduced stomatal conductance ( g S ) to improve water use efficiency (WUE) and, as a consequence, net photosynthetic rate ( P n ), transpiration rate ( T r ), and intercellular CO 2 concentration ( C i ) were also decreased in DS-R T .118 plants compared with C plants. These reductions became more significant with decreasing soil water availability. Correlation analysis showed g S was positively correlated ( P < 0.01) with the soil water content as well as leaf relative water content (RWC). There was a tendency to accumulate proline, malondialdehyde (MDA), antioxidases, and membrane electrolyte leakage as stress intensity increased. Moreover, drought stress induced significant ( P < 0.05) decline in total chlorophyll contents (Chl t ) and increase of nonphotochemical quenching (NPQ) on day 8 as a photo-protective mechanism. Cluster analysis distinguished the adaption of T .118 plants to water deficit in two ways. First, photosynthesis was related to thermal dissipation, and second antioxidation was related to morphology and osmosis. Furthermore, tested parameters showed a reversed tendency and restored equivalently to C levels after 9 days of rewatering. These findings suggest that T .118 plants demonstrated considerable tolerance to short-term drought stress and recovery due to a high degree of plasticity in physiological acclimation.

In this study, three different strains of Taxodium hybrid 'Zhongshanshan' varieties [T. hybrid 302 (T. distichum♀×T. mucronatum♂), T. hybrid 407 (T. mucronatum♀×T. distichum♂), T. hybrid 118 (T. hybrid 302 ♀×T. macronatum ♂)] and their parents, T. distichum and T. mucronatum, were applied to investigate the response of photosynthetic characteristics, antioxidant enzyme systems and morphological characteristics to drought stress and recovery. The results indicated that as drought days were prolonged, all plants' net photosynthetic rate (Pn) decreased, while proline accumulated. Meanwhile, the antioxidases functioned to eliminate malonaldehyde toxicity. On the 8th day, the decrease of Pn of T. distichum was the biggest, T. hybrid 118 plants showed the highest water use efficiency and the smallest MDA content, while T. macronatum plants increased the activity of superoxide dismutase and content of proline. After rewatering for 2 days, all these parameters showed signs of recovery, and the T. hybrid 118 plants showed the fastest recovery rate since their Pn and proline content had recovered for 74.4% and 60.2%, respectively. Then after recovered for 9 days, all tested parameters had almost restored to equivalent levels of CK plants. The total biomass of T. hybrid 118 plants was not affected, while the ratio of root to shoot was significantly (P<0.05) increased. The drought-resistance capacity ranged as T. macronatum>T. hybrid 118>T. hybrid 407>T. hybrid 302>T. distichum plants. In conclusion, the backcross generation T. hybrid 118 plants largely inherited the drought resistance of T. mucronatum, and the result would be instrumental in breeding and popularization of drought-resistant hybrid varieties.