Genome-Wide Identification and Expression Analysis of the bHLH Transcription Factor Family and Its Response to Abiotic Stress in Mongolian Oak (Quercus mongolica).

The basic helix-loop-helix (bHLH) family, one of the largest families of transcription factors in plants, is extensively involved in the growth, development, and stress response of several woody plants. However, no systematic analysis of the bHLH gene family in Quercus mongolica has been reported. We characterize QmbHLH genes and identify the functions of QmbHLH proteins in Q. mongolica. We used bioinformatics approaches, qRT-PCR analysis, and RNA sequencing data to examine chromosomal distributions, gene structures, and conserved patterns, and identified 89 QmbHLH genes, which were divided into 21 subgroups based on the phylogenetic analysis of bHLH genes in Arabidopsis thaliana. Segmental replication played a more prominent role than tandem duplication in the expansion of the QmbHLH gene family. Based on patterns of tissue-specific expression, protein interactions, and cis-element analysis, QmbHLH genes may be extensively involved in the growth and development of Q. mongolica. In leaves, stems, and roots, 12 selected QmbHLH genes exhibited responsiveness to abiotic stresses (salt, cold, weak light, and drought). Our study facilitates follow-up functional investigations of the bHLH gene family in Q. mongolica and provides novel insights into bHLH superfamilies in woody plants.

Identification of Functional Brassinosteroid Receptor Genes in Oaks and Functional Analysis of QmBRI1.

Brassinosteroids (BRs) play important regulatory roles in plant growth and development, with functional BR receptors being crucial for BR recognition or signaling. Although functional BR receptors have been extensively studied in herbaceous plants, they remain largely under-studied in forest tree species. In this study, nine BR receptors were identified in three representative oak species, of which BRI1s and BRL1s were functional BR receptors. Dispersed duplications were a driving force for oak BR receptor expansion, among which the Brassinosteroid-Insensitive-1 (BRI1)-type genes diverged evolutionarily from most rosids. In oak BRI1s, we identified that methionine in the conserved Asn-Gly-Ser-Met (NGSM) motif was replaced by isoleucine and that the amino acid mutation occurred after the divergence of Quercus and Fagus. Compared with QmBRL1, QmBRI1 was relatively highly expressed during BR-induced xylem differentiation and in young leaves, shoots, and the phloem and xylem of young stems of Quercus mongolica. Based on Arabidopsis complementation experiments, we proved the important role of QmBRI1 in oak growth and development, especially in vascular patterning and xylem differentiation. These findings serve as an important supplement to the findings of the structural, functional and evolutionary studies on functional BR receptors in woody plants and provide the first example of natural mutation occurring in the conserved BR-binding region (NGSM motif) of angiosperm BRI1s.

Group V Chitin Deacetylases Influence the Structure and Composition of the Midgut of Beet Armyworm, Spodoptera exigua.

Chitin deacetylase (CDA) can accelerate the conversion of chitin to chitosan, influencing the mechanical properties and permeability of the cuticle structures and the peritrophic membrane (PM) in insects. Putative Group V CDAs SeCDA6/7/8/9 (SeCDAs) were identified and characterized from beet armyworm Spodoptera exigua larvae. The cDNAs of SeCDAs contained open reading frames of 1164 bp, 1137 bp, 1158 bp and 1152 bp, respectively. The deduced protein sequences showed that SeCDAs are synthesized as preproteins of 387, 378, 385 and 383 amino acid residues, respectively. It was revealed via spatiotemporal expression analysis that SeCDAs were more abundant in the anterior region of the midgut. The SeCDAs were down-regulated after treatment with 20-hydroxyecdysone (20E). After treatment with a juvenile hormone analog (JHA), the expression of SeCDA6 and SeCDA8 was down-regulated; in contrast, the expression of SeCDA7 and SeCDA9 was up-regulated. After silencing SeCDAV (the conserved sequences of Group V CDAs) via RNA interference (RNAi), the layer of intestinal wall cells in the midgut became more compact and more evenly distributed. The vesicles in the midgut were small and more fragmented or disappeared after SeCDAs were silenced. Additionally, the PM structure was scarce, and the chitin microfilament structure was loose and chaotic. It was indicated in all of the above results that Group V CDAs are essential for the growth and structuring of the intestinal wall cell layer in the midgut of S. exigua. Additionally, the midgut tissue and the PM structure and composition were affected by Group V CDAs.

Identification and functional analysis of two potential RNAi targets for chitin degradation in Holotrichia parallela Motschulsky (Insecta Coleoptera).

Chitin metabolism enzymes are safe and desirable targets for pest management. ß-N-acetylglucosaminidase (NAG) and N-acetylglucosamine kinase (NAGK) are involved in chitin degradation. NAG is the main glycosidase that works synergistically with chitinases. NAGK is a key enzyme for the generation of UDP-Nacetylglucosamine (UDP-GlcNAc) and for the conversion of GlcNAc into GlcNAc 6-phosphate (GlcNAc-6-P). In this study, NAG and NAGK genes were identified from Holotrichia parallela, a polyphagous soil pest that causes serious damage to crops. The spatiotemporal expression investigated by RT-qPCR indicated that the two genes are expressed in all larval developmental stages. HpNAG is highly expressed in the integument and HpNAGK overexpressed in the midgut. After injection of dsHpNAG and dsHpNAGK, a significant RNAi effect was found after 72 h and larvae stopped growing. The survival rates of larvae were 13.3% and 16.7%, respectively. RNAi of HpNAG and HpNAGK regulated the expression levels of chitin metabolism-related genes, indicating that these two genes could be critical in the chitin metabolism. Furthermore, silencing HpNAG and HpNAGK reduced the thickness of the cuticle, and decreased its content of chitin. The study will lay a foundation for further clarifying the mechanism of chitin metabolism and provide potential targets for the biological control of H. parallela larvae.

RNA interference-mediated functional characterization of Group I chitin deacetylases in Holotrichia parallela Motschulsky.

Chitin deacetylases (CDAs, EC 3.5.1.41) catalyze the N-deacetylation of chitin to produce chitosan, which is essential for insect survival. Hence, CDAs are promising targets for the development of novel insecticidal drugs. In this study, the putative Group I chitin deacetylase genes HpCDA1, HpCDA2-1 and HpCDA2-2 were identified from Holotrichia parallela. Conserved domain database search identified a chitin-binding peritrophin-A domain (ChBD), a low-density lipoprotein receptor class A domain (LDLa), and a putative CDA-like catalytic domain. RT-qPCR analysis showed that the Group I HpCDAs were expressed in various tissues and predominant in the integument. The developmental expression patterns from the first-instar to third-instar larvae showed that HpCDAs were highly expressed on the first day and gradually declined after molting. The functional characteristics of the Group I CDAs in cuticle organization were examined using RNA interference (RNAi) and transmission electron microscopy (TEM) methods. Administration of double-stranded HpCDA (dsHpCDA) through larval injection could suppress the expression levels of HpCDA1 and HpCDA2, thus resulting in abnormal or lethal phenotypes. TEM analysis revealed that RNAi of either HpCDA1 or HpCDA2 remarkably affected the cuticle integrity, as evidenced by cuticle disorganization and chitin laminae disruption, suggesting the crucial role of CDAs in chitin modification. These experimental results demonstrate the important contribution of putative key genes involved in chitin metabolism, and provide a foundation for developing new strategies to control H. parallela.

Functions of microRNA in seed development, dormancy and germination processes.

In plants, microRNAs (miRNAs) are a class of non-coding small RNAs that play important roles in various biological processes, such as seed development, dormancy and germination. In these stages, seeds are regulated by various extrinsic environmental factors and intrinsic hormones. Different species of seeds have distinct development and dormancy characteristics. Recent investigations have identified more and more miRNAs to regulate seed morphogenesis, material metabolism and stress response capacities through hormonal signal transduction, antioxidant effects and transcription factors, etc. In this review, we summarize recent progresses on regulatory mechanisms of miRNAs, and also complex multilevel regulation of miRNAs in the process of seed development, dormancy and germination. We hope to provide insights into further research.

Identification of a new peritrophic membrane protein from larval Holotrichia parallela (Coleoptera: Motschulsky).

Peritrophic membranes (PMs) are composed of proteins, proteoglycans and chitin that play important roles in the structural formation and function of the PM. This study identified and characterized a new chitin binding protein named HpCBP45 by immunoscreening of the Holotrichia parallela larvae midgut expression library. The predicted amino acid sequence indicates that it contains eight tandem chitin binding domains belonging to the peritrophin-A family. The HpCBP45 protein was expressed as a recombinant protein in the yeast Pichia pastoris and chitin binding assay demonstrated that recombinant HpCBP45 protein could strongly bind to chitin. qRT-PCR analysis showed that HpCBP45 was mainly localized in the midgut, further confirming the H. parallela PM belongs to Type I PM. The discovery and characterization of the peritrophic membrane protein HpCBP45 provides a basis for the further investigation of its biochemical and physiological functions in H. parallela.

Identification and Functional Analysis of V-ATPaseA and C Genes in Hyphantria cunea.

Vacuolar (H+)-ATPases (V-ATPases) are ATP-driven proton pumps that play multifaceted roles across various organisms. Despite their widespread significance, the functional implications of V-ATPase genes in Hyphantria cunea, an invasive forest pest with a global presence, have yet to be elucidated. In this study, two specific V-ATPase genes from H. cunea were identified and analyzed, namely HcV-ATPase A (accession number: OR217451) and HcV-ATPase C (accession number: OR217452). Phylogenetic analysis and multiple sequence alignment reveal that HcV-ATPase A shares the highest amino acid sequence similarity with SfV-ATPase A, while HcV-ATPase C is most similar to HaV-ATPase C. Spatiotemporal expression profiles, determined via RT-qPCR, demonstrate that both HcV-ATPase A and HcV-ATPase C are expressed throughout all larval developmental stages, with HcV-ATPase A predominantly expressed in the midgut and HcV-ATPase C showing high expression in the epidermis. RNA interference (RNAi) targeting of these genes significantly suppressed their expression by 62.7% and 71.0% 120 h post-injection, leading to halted larval growth and increased mortality rates of 61.7% and 46.7%, respectively. Further investigations using immunohistochemistry, hematoxylin and eosin (HE) staining, and transmission electron microscopy (TEM) revealed that gene silencing induced vesiculation and subsequent losses or sloughing of intestinal parietal cells, alongside an increase in the number of autophagic cells. Additionally, the silencing of HcV-ATPase A and C genes resulted in a reduced gut epidermal cell layer thickness and further increases in goblet cell numbers. Importantly, RNAi of HcV-ATPase A and C did not affect the expression levels of one another, suggesting independent functional pathways. This study provides foundational insights into the role of V-ATPase in H. cunea and identifies potential targets for the biocontrol of its larvae, contributing to the understanding of V-ATPase mechanisms and their application in pest management strategies.

Novel Environmentally Friendly RNAi Biopesticides: Targeting V-ATPase in Holotrichia parallela Larvae Using Layered Double Hydroxide Nanocomplexes.

RNA interference (RNAi)-based biopesticides offer an attractive avenue for pest control. Previous studies revealed high RNAi sensitivity in Holotrichia parallela larvae, showcasing its potential for grub control. In this study, we aimed to develop an environmentally friendly RNAi method for H. parallela larvae. The double-stranded RNA (dsRNA) of the V-ATPase-a gene (HpVAA) was loaded onto layered double hydroxide (LDH). The dsRNA/LDH nanocomplex exhibited increased environmental stability, and we investigated the absorption rate and permeability of dsRNA-nanoparticle complexes and explored the RNAi controlling effect. Silencing the HpVAA gene was found to darken the epidermis of H. parallela larvae, with growth cessation or death or mortality, disrupting the epidermis and midgut structure. Quantitative reverse transcription-polymerase chain reaction and confocal microscopy confirmed the effective absorption of the dsRNA/LDH nanocomplex by peanut plants, with distribution in roots, stems, and leaves. Nanomaterial-mediated RNAi silenced the target genes, leading to the death of pests. Therefore, these findings indicate the successful application of the nanomaterial-mediated RNAi system for underground pests, thus establishing a theoretical foundation for developing a green, safe, and efficient pest control strategy.

Transcriptomic insights into the effects of abscisic acid on the germination of Magnolia sieboldii K. Koch seed.

Magnolia sieboldii K. Koch is a deciduous tree species. However, the wild resource of M. sieboldii has been declining due to excessive utilization and seed dormancy. In our previous research, M. sieboldii seeds have morphophysiological dormancy and low germination rates under natural conditions. The aim of the present study was to identify the genes involved in dormancy maintenance. In this study, the germination percentage of M. sieboldii seeds negatively correlated with the content of endogenous abscisic acid (ABA). The hydration of seeds for germination showed three distinct phases. Five key time points were identified: 0 h imbibition (dry seed, GZ), 0 day after imbibition (DAI), 16 DAI, 40 DAI, and 56 DAI. The comprehensive transcript profiles of M. sieboldii seeds treated with ABA and water at the five key germinating stages were obtained. A total of 9641 differentially expressed genes (DEGs) were identified, and 208 and 197 common DEGs were found throughout the ABA and water treatments, respectively. Compared with that in the GZ, 518, 696, 2133, and 1535 DEGs were identified in the SH group at 0, 16, 40 and 56 DAI, respectively. 666, 1725, 1560 and 1415 DEGs were identified in the ABA group at 0, 16, 40, and 56 DAI, respectively. Among the identified DEGs, 12 722 were annotated with GO terms, the top three enriched GO terms were different among the DEGs at 56 DAI in the ABA vs. SH treatments. KEGG pathway enrichment analysis for DEGs indicated that oxidative phosphorylation, protein processing in endoplasmic reticulum, starch and sucrose metabolism play an important role in seed response to ABA. 1926 TFs are obtained and classified into 72 families from the M. sieboldii transcriptome. Results of differential gene expression analysis together with qRT-PCR indicated that phase II is crucial for rapid and successful seed germination. This study is the first to present the global expression patterns of ABA-regulated transcripts in M. sieboldii seeds at different germinating phases.

Modeling knot features using branch scars from Mongolian oak (Quercus mongolica).

Wood quality is an important indicator for modern sawmills. Internal wood characteristics can be derived from their correlations with external appearances. In this study, we developed linear regression models to predict knot size from surface features of Mongolian oak (Quercus mongolica) using data collected from 53 trees. For this, manual measurements and X-ray computed tomography scanning technology was respectively used to obtain internal and external features of 1,297 knots. Our results showed that Mongolian oak knots were generally concentrated in the middle part of oak stems, with fewer knots observed at the top and base. The parameters of knot and scar showed significant correlations (P < 0.01), where length and diameter of the corresponding external scar increase with increasing the length and diameter of a knot. The corresponding external scar can be used as an effective indicator to predict the internal value of oak logs. The accuracy of our constructed model is more than 95% when assessed against independent test samples. These models thus can be applied to improve the practical production of oak timber and reduce commercial loss caused by knots. These additional data can improve the estimation of the influence of knots on wood quality and provide a theoretical foundation for investigating the characteristics of hardwood knots.

Composition and Diversity of Endophytic Rhizosphere Microbiota in Apple Tree with Different Ages.

In order to determine the underlying mechanism of the senescence occurring in older apple trees, the effects of tree age on the community structure and dominant genus of endophytic rhizosphere bacteria in apple were investigated. The diversity and structure of the bacterial communities and corresponding changes in the dominant genera of endophytic rhizosphere bacteria of apple at different ages (2, 8, 16, 22 years) were compared based on 16S rRNA high-throughput sequencing technology. The results revealed that the longer the tree age, the less the number of ASV in the endophytic bacteria. Moreover, the number of ASV in the endophytic bacteria gradually decreased as the tree age increased, however no significant changes were observed in the alpha diversity. At the phyla level, the relative abundance of Actinobacteria increased, while that of Proteobateria decreased. At the genus level, the relative abundance of Mycobacterium, Chujaibacter, and other genera increased, while the relative abundance of Aquabacterium, Ralstonia, Streptomyces, Asticcacaulis, Hyphomicrobium, Pseudomonas, and Sphingomonas decreased. The reduced relative abundance of endophytic rhizosphere bacteria associated with plant growth and disease resistance may thus be the cause of tree senescence. This work acts as a reference to increases the understanding of plant-microbe interactions.

Genome-wide identification of the auxin response factor (ARF) gene family in Magnolia sieboldii and functional analysis of MsARF5.

Auxin plays an essential role in flowering, embryonic development, seed dormancy, and germination. Auxin response factors (ARFs) are plant-specific key transcriptional factors in mediating the gene expression network of auxin signaling. Although ARFs in model plants such as Arabidopsis had been well characterized, their identities and potential roles in non-model plants are less studied. Here, we performed genome-wide identification of ARFs in Magnolia sieboldii K. Koch, a primitive species with high taxonomic importance and medicinal values. We found 25 ARF genes in M. sieboldii, which were widely distributed across multiple chromosomes. Based on sequence similarity, the encoded proteins could be either transcriptional repressors or activators. Gene expression analysis showed a dynamic pattern for many ARFs including MsARF5 during seed germination. In addition, overexpressing of MsARF5 showed that it restores many developmental defects in the Arabidopsis mutant. Moreover, two phenotypically distinct transgenic Arabidopsis lines were obtained, indicating a link between gene expression levels and developmental phenotypes. Taken together, we provided a systematic investigation of the ARF gene family in M. sieboldii and revealed an important role of MsARF5 in mediating auxin signaling.

Selection and validation of reference genes for quantitative real-time PCR of Quercus mongolica Fisch. ex Ledeb under abiotic stresses.

Quercus mongolica Fisch. ex Ledeb is the main species of coniferous and broadleaved mixed forests in northeast and north China, which has high ornamental, economic, and ecological value. The appropriate reference genes must be selected for quantitative real-time PCR to reveal the molecular mechanisms of stress responses and their contribution to breeding of Q. mongolica. In the present study, we chose 11 candidate reference genes (TUA, CYP18, HIS4, RPS13, ACT97, TUB1, UBQ10, UBC5, SAND, PP2A, and SAMDC) and used four programs (GeNorm, NormFinder, BestKeeper, and RefFinder) to assess the expression stability of the above genes in roots, stems, and leaves under five abiotic stress factors (cold, salt, drought, weak light, and heavy metal). The findings revealed that under various experimental environments, the most stable genes were different; CYP18, ACT97, and RPS13 ranked the highest under most experimental environments. Moreover, two genes induced by stress, CMO and P5CS2, were chosen to demonstrate the reliability of the selected reference genes in various tissues under various stress conditions. Our research provides a significant basis for subsequent gene function studies of Q. mongolica.

[Responses of stand growth, regeneration, and understory species diversity in Quercus mongolica secondary forest to stand density.] / è’™å¤æ Žæ¬¡ç”Ÿæž—çš„ç”Ÿé•¿æ›´æ–°ä¸Žæž—ä¸‹æ¤è¢«å¤šæ ·æ€§å¯¹æž—åˆ†å¯†åº¦çš„å“åº”.

Six Quercus mongolica plots with an area of 0.1 hm2 were thinned in 2018. A field survey was carried out in 2020 to examine the effects of different stand densities (high: 900 trees·hm-2; medium: 720 trees·hm-2; low: 600 trees·hm-2) on growth and regeneration of stands and understory species diversity of secondary Q. mongolica forests in Qingyuan, Liaoning Province. Due to the short interval after thinning, there was no significant difference in tree height and diameter at breast height under different densities. However, the crown symmetry index under low stand density was significantly higher than that of high stand density, indicating that crown growth was more sensitive to stand density than trunk growth. The abundance of seedlings was the highest in the medium density, and the basal diameter of the seedlings with the same height was significantly higher, and the seedling regeneration and growth at the medium density were much better than the other two densities. A total of 70 species were recorded, belonging to 41 families and 67 genera. Quercus mongolica, Lespedeza bicolor, Melampyrum roseum, and Potentilla freyniana were the dominant species of trees and herbs, respectively. Simpson index, Pielou index and Shannon index of shrub layer and herb layer were the highest at the medium density. It indicated that the stand density of 720 trees·hm-2 could help maintain the sustainable development of Q. mongolica secondary forest in the mountainous area of eastern Liaoning.

A chromosome-scale genome assembly of the Mongolian oak (Quercus mongolica).

Mongolian oak (Quercus mongolica Fisch.) is an ecologically and economically important white oak species native to and widespread in the temperate zone of East Asia. Here, we present a chromosome-scale reference genome assembly of Q. mongolica, a representative white oak species, by combining Illumina and PacBio data with Hi-C mapping technologies that is the first reference genome created for an Asian oak. Our results showed that the PacBio draft genome size was 809.84 Mb, with a BUSCO complete gene percentage of 92.71%. Hi-C scaffolding anchored 774.59 Mb contigs (95.65% of draft assembly) onto 12 pseudochromosomes. The contig N50 and scaffold N50 were 2.64 and 66.74 Mb, respectively. Of the 36,553 protein-coding genes predicted in the study, approximately 95% had functional annotations in public databases. A total of 435.34 Mb (53.75% of the genome) of repetitive sequences were predicted in the assembled genome. Genome evolution analysis showed that Q. mongolica is closely related to Q. robur from Europe, and they shared a common ancestor ~11.8 million years ago (Ma). Gene family evolution analysis of Q. mongolica revealed that the nucleotide-binding site (NBS)-encoding gene family related to disease resistance was significantly contracted, whereas the ECERIFERUM 1 (CER1) homologous genes related to cuticular wax biosynthesis was significantly expanded. This pioneering Asian oak genome resource represents an important supplement to the oak genomics community and will improve our understanding of Asian white oak biology and evolution.

Integrated RNA and miRNA sequencing analysis reveals a complex regulatory network of Magnolia sieboldii seed germination.

Magnolia sieboldii K. Koch (M. sieboldii) is a deciduous Chinese tree species of the Magnoliaceae family with high ornamental, medicinal, and economic benefits. The germination of M. sieboldii seeds under natural conditions is extremely difficult, thereby hindering the cultivation and breeding of this important species. The molecular mechanisms underlying M. sieboldii seed germination remain unclear due to the lack of genomic and transcriptomic resources. Here, we integrated both mRNA and miRNA sequencing to identify the genes and pathways related to M. sieboldii germination. A comprehensive full-length transcriptome containing 158,083 high-quality unigenes was obtained by single-molecule real-time (SMRT) sequencing technology. We identified a total of 13,877 genes that were differentially expressed between non-germinated and germinated seeds. These genes were mainly involved in plant hormone signal transduction and diverse metabolic pathways such as those involving lipids, sugars, and amino acids. Our results also identified a complex regulatory network between miRNAs and their target genes. Taken together, we present the first transcriptome of M. sieboldii and provide key genes and pathways associated with seed germination for further characterization. Future studies of the molecular basis of seed germination will facilitate the genetic improvement M. sieboldii.

Transcriptome-based analysis of the hormone regulation mechanism of gender differentiation in Juglans mandshurica Maxim.

Juglans mandshurica Maxim is a hermaphroditic plant belonging to the genus Juglans in the family Juglandaceae. The pollination period of female flowers is different from the loose powder period of male flowers on the same tree. In several trees, female flowers bloom first, whereas in others, male flowers bloom first. In this study, male and female flower buds of J. mandshurica at the physiological differentiation stage were used. Illumina-based transcriptome sequencing was performed, and the quality of the sequencing results was evaluated and analyzed. A total of 138,138 unigenes with an average length of 788 bp were obtained. There were 8,116 differentially expressed genes (DEGs); 2,840 genes were upregulated, and 5,276 genes were downregulated. The DEGs were classified by Gene Ontology and analyzed by Kyoto Encyclopedia of Genes and Genomes. The signal transduction factors involved in phytohormone synthesis were selected. The results displayed that ARF and SAUR were expressed differently in the auxin signaling pathway. Additionally, DELLA protein (a negative regulator of gibberellin), the cytokinin synthesis pathway, and A-ARR were downregulated. On April 2nd, the contents of IAA, GA, CTK, ETH and SA in male and female flower buds of two types of J. mandshurica were opposite, and there were obvious genes regulating gender differentiation. Overall, we found that the sex differentiation of J. mandshurica was related to various hormone signal transduction pathways, and hormone signal transduction plays a leading role in regulation.

Flower Development of Heterodichogamous Juglans mandshurica (Juglandaceae).

Juglans mandshurica is a monoecious heterodichogamous species with protogynous and protandrous mating strategies that occur at a 1:1 ratio and are randomly distributed in the population. The inconsistent male and female flowering periods of the same mating type result in an imbalance of the ratio of male and female flowers, contributing to the low yield of this species. However, little more is known about its floral development. Following three consecutive years of observations, histological analysis, and scanning electron microscopy, we found that the morphological and anatomical development of the male and female flowers were synchronous. The male floral morphological development of J. mandshurica was divided into seven phases, while that of the female flower was nine. Four stages were shared between the male and female flower's anatomical development. Our findings indicate that there was minimal overlap between sexual functions within the same mating type, guaranteeing synchronization, mutual non-interference, outcrossing, and avoidance of self-fertilization. These results provide a theoretical basis for the improvement of fruit yield and quality through the reasonable allocation of protogynous and protandrous individuals in a population, and for artificial pollination control. Further, these findings lay a foundation for further research on the genetic mechanisms and environmental effects on flower development of heterodichogamous J. mandshurica.

Cloning and expression analyses of a Pyrabactin Resistance 1 (PYR1) gene from Magnolia sieboldii K. Koch.

Magnolia sieboldii K. Koch is endemic to China and has high medicinal and ornamental values. However, its seed exhibits morphophysiological dormancy, and the molecular mechanisms of which are not clearly understood. To reveal the regulation mechanism of the ABA signal in seed dormancy, the M. sieboldii ABA receptor Pyrabactin Resistance 1 (PYR1) gene was cloned and analyzed. Analysis of the MsPYR1 sequence analysis showed that the full-length cDNA contained a complete open reading frame of 987 bp and encoded a predicted protein of 204 amino acid residues. The protein had a relative molecular weight of 22.661 kDa and theoretical isoelectric point of 5.01. The transcript levels of MsPYR1 were immediately upregulated at 16 DAI and then decreased at 40 DAI. The highest transcript level of MsPYR1 was found in the dry seeds, indicating that the MsPYR1 gene may play an important role in the regulation of dormancy. The MsPYR1 gene cDNA was successfully expressed in E. coli Rosetta (DE3), and the protein bands were consistent with the prediction. The Anti-MsPYR1antibody could detect the expression of MsPYR1 in M. sieboldii. The results provided a foundation for further study of the function of the MsPYR1 gene.ABBREVIATIONSABA: Abscisic acid; MPD: morphophysiological; PYR1: Pyrabactin Resistance1; PYL: Pyr1-Like; RCAR: Regulatory Components of Aba Receptors; PP2C: protein phosphatases 2C; SnRK2: sucrose non-fermenting1-related protein kinase2; DAI: day after imbibition; NCBI: National Center for Biotechnology Information; BCA: Bicinchoninic acid; CDD: Conserved Domains.