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    Gene Cloning, Expression and Interaction Protein Analysis of FERONIA in Brassica oleracea var. acephala
    Baoru XUN, Hongtao QIN, Rui MA, Nanfeng GUO, Yunping LIU, Ying WU, Xingguo LAN
    Bulletin of Botanical Research    2024, 44 (2): 298-306.   DOI: 10.7525/j.issn.1673-5102.2024.02.015
    Abstract60)   HTML4)    PDF(pc) (4370KB)(57)       Save

    FERONIA (FER) receptor kinase plays an important role in pollen and stigma recognition in flowering plants. To analyze the function of receptor-like kinase FERONIA(FER) in the pollination in ornamental kale(Brassica oleracea var. acephala), BoFER gene was cloned from the stigma of the self-incompatibility line(S13-bS13-b ) of ornamental kale by RT-PCR, and the cloned cDNA sequence was 2 682 bp, which encoded 893 amino acids containing the highly conserved kinase domain and Ser/Thr kinase binding site, and the expression level of BoFER was analyzed by qRT-PCR. The results showed that the expression of BoFER in stigmas gradually increased after incompatible pollination, but gradually decreased after compatible pollination. Furthermore, the interaction between BoFER and known proteins related to self-incompatibility were analyzed by yeast two-hybrid. The results suggested that BoFER interacted with the kinase domain of S-locus receptor kinase BoSRK13-b.

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    Construction and Identification of TRV-mediated VIGS Transformation System of Iris sanguinea
    Guiling LIU, Nuo XU, Gongfa SHI, Ling WANG
    Bulletin of Botanical Research    2024, 44 (1): 132-138.   DOI: 10.7525/j.issn.1673-5102.2024.01.015
    Abstract127)   HTML5)    PDF(pc) (1718KB)(229)       Save

    To analyze gene function in plants lacking stable genetic transformation system, virus-induced gene silencing(VIGS) was needed, and Iris sanguinea, a monocotyledon, was selected as materials. The specific fragment of IsPDS gene was isolated and the VIGS recombinant vector pTRV2-IsPDS was constructed and leaves were infected by injection. The results showed that the most effective infection was achieved by injecting its leaf veins with syringes when the OD600 values of the resuspension were adjusted to 0.8-1.0 after the those of pTRV1 and pTRV2-IsPDS were adjusted to 1.8-2.0. The experiment was conducted when outdoors temperature was 15-20 ℃ from 6 p.m. to 8 p.m., and a 1 mL syringe needle pricking the outer epidermis of I. sanguinea leaves and 1 mL of heavy suspension slowly injected along parallel veins into its vascular bundles. A clear albino phenotype might appear after about 14 days. TRV1 and TRV2 virus vectors were detected in the plants with phenotypic changes and in the no-load group. The expression of IsPDS in the albino plants was significantly lower than that in the no-load group and the control group. With the concentration of agrobacterium tumefaciens carrying virus vector increased in the preparation of infection solution, the infection efficiency of the whole experiment was improved, and no shading was needed after inoculation.

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    Bioinformatics Analysis of CbuDELLAs Gene Family and Functional Analysis of CbuGRAS9
    Shanshan WANG, Rui WANG, Erqin FAN, Pengyue FU, Guanzheng QU, Nan WANG
    Bulletin of Botanical Research    2024, 44 (1): 139-151.   DOI: 10.7525/j.issn.1673-5102.2024.01.016
    Abstract56)   HTML6)    PDF(pc) (6650KB)(117)       Save

    To provide theoretical basis for genetic improvement of reproductive regulatory traits of Catalpa bungei, DELLAs family genes were identified and the function of CbuGRAS9 was analyzed. Based on the genomic data of Catalpa bungei, five CbuDELLAs genes homologous to Arabidopsis thaliana were identified and cloned. ExPASy, SWISS-MODEL, Plant-mPloc, PlantCare and other online tools were used to predict the isoelectric point, protein structure, sub-cellular localization and promoter cis-acting elements of CbuDELLAs protein. The expression differences of CbuDELLAs were analyzed by using Catalpa ‘Bairihua’ and Catalpa bungei ‘Luoqiu No.1’ as materials, and the molecular function of CbuGRAS9 was confirmed by heterologus transformation in Arabidopsis thaliana, and the proteins interacted with CbuGRAS9 were screened by yeast two-hybrid library. The results showed that the amino acid number of the five CbuDELLAs proteins ranged from 455 to 588 aa, the relative molecular weight of the proteins ranged from 5.04 to 6.43 kDa, and the isoelectric point value ranged from 4.81 to 5.14. All CbuDELLAs proteins contained DELLA and GRAS conserved domains and are hydrophilic proteins. Sub-cellular localization prediction showed that CbuDELLAs protein was located in the nucleus. The analysis of promoter cis-acting elements showed that the five promoter regions of DELLAs all contained cis-acting elements involved in gibberellin reaction. The results of qRT-PCR showed that the expression of CbuDELLAs in Catalpa ‘Bairihua’ were significantly higher than that in Catalpa bungei ‘Luoqiu No.1’, and CbuGRAS9 was the most significantly gene, and the flowering time of CbuGRAS9 transgenic plants were significantly delayed. Proteins interacted with CbuGRAS9 were mainly concentrated in metabolic pathways such as ribosome, amino acid synthesis, secondary metabolism, photosynthesis and TCA cycle.

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    Expression Pattern Analysis of PdbHMGs Genes in Populus davidiana×P. bolleana under Abiotic Stress
    Chunyao WANG, Xiaojin LEI, Zhongyuan LIU
    Bulletin of Botanical Research    2023, 43 (6): 932-942.   DOI: 10.7525/j.issn.1673-5102.2023.06.015
    Abstract51)   HTML8)    PDF(pc) (3486KB)(109)       Save

    In order to investigate the function of HMG(High mobility group proteins) genes under abiotic stresses in Populus davidiana×P. bolleana, the HMG genes were cloned and the stress tolerant function of HMGs was identified, and the theoretical foundation for using the gene in tree breeding through genetic engineering was laid. Seven homologous genes of ArabidopsisHMGs(named PdbHMG1-7) were found by homologous sequence alignment, and sequence analysis of PdbHMG1-7 was performed using bioinformatics tools respectively, the expression patterns of PdbHMG1-7 gene in leaves and roots of Populus davidiana×P.bolleana under different stresses were investigated by qRT-PCR.Results The ORF length of PdbHMG1-7 were 456-2 004 bp respectively, the amino acid residues encoded by PdbHMG1-7 genes were 151-667 aa, and the molecular weight of the encoded protein were 16.72-75.54 kDa, and the theoretical isoelectric point were 5.35-9.36, and the seven PdbHMG were hydrophilic proteins. The results of multiple sequence alignment and phylogenetic tree analysis showed that all 7 PdbHMG genes contained HMG-box conserved domains and belonged to HMGB subfamily. The analysis results of cis-acting elements contained in the promoter showed that the promoter sequence of PdbHMG genes contained a variety of elements in response to hormones and stress. The expression pattern analysis showed that the expression levels of seven PdbHMGs in roots and leaves under NaCl, PEG6000, ABA and CdCl2 stress treatment changed in at least one stress treatment time point respectively. In leaves, PdbHMG2 were induced under the stress of NaCl, PEG6000, ABA and CdCl2, and almost all of them were significantly up-regulated respectively, but PdbHMG7 was down-regulated. Under NaCl and CdCl2 stress, PdbHMG1PdbHMG3 and PdbHMG6 were almost significantly up-regulated during the whole stress process respectively. The expressions of PdbHMG3 in leaves were also significantly up-regulated at most time points under NaCl and CdCl2 stress. In roots, the expression of PdbHMG3 and PdbHMG5 was significantly up-regulated in the whole process of NaCl treatment. However, PdbHMG6 was not significantly expressed during NaCl stress except for 9-12 h, the expression of PdbHMG6 was significantly down-regulated at other stress time points. During ABA treatment, PdbHMG6 was down-regulated in roots. Conclusion The results showed that seven PdbHMG genes could respond to at least one stress treatment in one organ, but the types and mechanisms of involved stress response might be different. This study provided a reference for further studying the function of PdbHMGs family genes in Populus davidiana×P. bolleana.

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    Cloning and Expression Analysis of Catalase Gene in Lilium pumilum
    Yu SONG, Wenhao LIN, Yibo JING, Yi DONG, Shumei JIN
    Bulletin of Botanical Research    2023, 43 (5): 756-767.   DOI: 10.7525/j.issn.1673-5102.2023.05.012
    Abstract103)   HTML9)    PDF(pc) (6823KB)(97)       Save

    To explore the relationship between the CatalaseCAT) gene and saline-alkali stress tolerance of Lilium pumilum, the CAT gene was successfully cloned from L. pumilum bulb. The length of region ORF was 1 479 bp, encoding 492 amino acids, and sequence alignment and phylogenetic tree analysis were performed, and the Catalase protein of L. pumilum was found to be closely related to the Catalase proteins of L. sargentiaeAnanas comosusElaeis guineensis and other plants. The Catalase protein was induced to express, and purified in vitro with 1 mol·L-1 IPTG as the inducer after constructing prokaryotic expression vector pQE-LpCat. The results showed that the growth concentration of bacteria solution containing PQE-LpCat protein was higher than that of the control strain under the stress of 50 mmol·L-1 NaHCO3. Under the stress of 1 mol·L-1 NaHCO3 and 2.5 mol·L-1 H2O2, less wilting was observed in plants overexpressing the LpCAT gene compared to wild-type. Determination of the physiological indexes including net photosynthetic rate, stomatal conductance, intercellular CO2 and transpiration rate, H2O2 concentration and contents of malondialdehyde(MDA) showed that tobacco plants overexpressing the LpCat gene were more tolerant to saline and alkali stress than wild-type tobacco plants.

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    Functional Analysis of FmCCoAOMT Gene in Fraxinus mandshurica During Lignin Synthesis and Abiotic Stress
    Chaoran SHAN, Xiaohu CHEN, Yunfei DING, Wei ZHAO, Han LU, Shangzhu GAO, Fenghui QI, Yaguang ZHAN, Fansuo ZENG
    Bulletin of Botanical Research    2023, 43 (5): 768-778.   DOI: 10.7525/j.issn.1673-5102.2023.05.013
    Abstract1284)   HTML9)    PDF(pc) (3077KB)(113)       Save

    To clarify the function of FmCCoAOMT gene, the expression pattern of FmCCoAOMT gene was analyzed by qRT-PCR using Fraxinus mandshurica as material in this study. The results showed that FmCCoAOMT gene was tissue specific. The expression level was the highest in the xylem of both female and male perennial plants, and was 9.4 times higher than that in the bark of new shoots in male plants, and was 13.1 times higher than that of leaf in female plants. The leaf disk method was used to obtain transgenic tobacco plants. Compared with the wild type, the lignin content of overexpressing FmCCoAOMT was increased by 41%, while the cellulose and hemicellulose contents were decreased by 32% and 52%, respectively. Tobacco overexpressing FmCCoAOMT after abiotic stress treatment compared to wild type, the activity of POD and SOD increased, the content of MDA decreased, and the content of reactive oxygen decreased after stress treatment. After NaCl treatment, POD activity was 83% higher than that of control group. After NaCl, ABA and mannitol treatments, SOD activity increased by 56.6%, 44.2% and 19.0%, and MDA content decreased by 77.2%, 11.7% and 47.6%, respectively. The results indicated that FmCCoAOMT gene might be involved in lignin synthesis of F. mandshurica, promoted abiotic stress tolerance.

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    Effect of Expression of Aeβ- AS gene from Aralia elata on the Content of Saponins in Nicotiana tabacum L.
    Qingqing HUO, Yuxin XIA, Jiale LI, Wei HAN, Shuya ZHANG, Zhe ZHANG, Meiling XIA, Wenhua GUO, Xiangling YOU
    Bulletin of Botanical Research    2023, 43 (5): 779-786.   DOI: 10.7525/j.issn.1673-5102.2023.05.014
    Abstract1409)   HTML3)    PDF(pc) (1728KB)(250)       Save

    To investigate the effect of β-amyrin synthase gene(Aeβ-AS) on triterpenoid saponin synthesis of Aralia elata, the Aeβ-AS gene was cloned and genetically transformed into tobacco. The expression difference of Aeβ-AS gene in different parts of transgenic tobacco was analyzed, and the expression levels of Aeβ-AS gene, upstream and downstream key enzyme genes and total triterpene content were detected respectively. The results showed that the plant overexpression vector PROKⅡ-Aeβ-AS was successfully constructed and transferred into wild type tobacco, and seven transgenic lines were obtained and expressed at the mRNA level, and the expression level in leaves was higher than that in roots and stems. In transgenic tobacco, the expression levels of Aeβ-AS gene and its upstream and downstream key enzyme genes were higher than those of wild type, and the relative expression levels of NtFPS and NtSS genes of strain L21 were the highest, and the relative expression levels of NtSE and Aeβ-AS genes of strain L30 were the highest. Compared with wild-type tobacco, the total triterpene content of GM tobacco was significantly increased(1.1-1.6 times). The results showed that synthesis of Aeβ-AS gene and heterologous transformation in tobacco could significantly increase the total triterpene content in transgenic tobacco.

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    Cloning and Functional Analysis of EgrWAT1 Gene in Eucalyptus grandis
    Haonan ZHANG, Shanshan CHEN, Jianmin XU, Ping LUO, Xiaoping WANG, Zhiru XU, Chunjie FAN
    Bulletin of Botanical Research    2023, 43 (4): 601-611.   DOI: 10.7525/j.issn.1673-5102.2023.04.013
    Abstract1549)   HTML3)    PDF(pc) (2728KB)(497)       Save

    In order to explore the role of WALLS ARE THIN(WAT1) in wood formation and response to stress in woody plants, bioinformatics tools was used for analysis, and quantitative Real-time PCR(qRT-PCR) was used to investigate the expression patterns of EgrWAT1L and EgrWAT1S in different tissues, internodes and in response to different stresses, and the gene EgrWAT1S and its other transcript EgrWAT1L were cloned from Eucalyptus grandis. The results showed that the EgrWAT1S was highly expressed in phloem, while EgrWAT1L was mainly expressed in roots, and the expression patterns were significantly different under methyl jasmonate(MeJA) and salicylic acid(SA) treatment, salt stress, phosphorus(P) and boron(B) deficiency, and even opposite under MeJA and SA. These results suggested that EgrWAT1L might affect EgrWAT1S expression through transcriptional regulation and further protein translation in response to hormone and stress treatments. The studies provided a basis for further elucidate the function in the growth and development of EgrWAT1 gene and also provided a possibility for future molecular breeding of Eucalyptus.

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    The Function of Salt and Alkaline Tolerance of WRKY42 Gene in Amorpha fruticosa
    Yu SUN, Yiteng ZHANG, Huihui CHENG
    Bulletin of Botanical Research    2023, 43 (4): 612-621.   DOI: 10.7525/j.issn.1673-5102.2023.04.014
    Abstract1997)   HTML11)    PDF(pc) (3957KB)(1491)       Save

    To investigate the role of WRKY transcription factors(TFs) in sensing saline-alkali stress signals and maintaining their tolerance function through physiological and biochemical regulatory pathways, and the WRKY42 gene of Amorpha fruticosa was cloned and the expression pattern in response to salt(NaCl) and alkali(NaHCO3) stress and tissue organs was analyzed, and the salinity tolerance function was also studied its by overexpression in tobacco. In this study, the AfWRKY42 gene was cloned based on transcriptome sequencing data of A. fruticosa L. under stress. Bioinformatics analysis showed that AfWRKY42 contained a WRKY structural domain, two low-complexity regions and a helix region. Phylogenetic tree analysis of amino acids revealed that AfWRKY42 was most closely related to WRKY47 of Cajanus cajan and WRKY42 of Mucuna pruriens. The localization of AfWRKY42 protein in mesophyll protoplasts of Arabidopsis thaliana was confirmed in nucleus by a transient gene expression system. Quantitative analysis of AfWRKY42 gene showed the highest expression in the shoot epidermis of Sophora japonica. Detection of expression patterns in roots and leaves treated with NaCl and NaHCO3 showed an overall increasing trend induced by it, suggesting that overall stress induced an increase in AfWRKY42 gene expression, and AfWRKY42 gene was associated with the regulation of salinity tolerance in plants. Analysis of salinity tolerance in 35S-initiated overexpressing T3 generation of tobacco lines transgenic for AfWRKY42 gene showed that the transgenic tobacco lines showed increased resistance after salinity stress treatment, it had higher chlorophyll and electrical conductivity and significantly lower malondialdehyde content than the wild type, indicating that AfWRKY42 played an important regulatory role in response to salinity stress. It would provide a WRKY transcription factor candidate gene for salinity resistance breeding and lay the foundation for improving the resistance of A. fruticosa and other plants.

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    Cloning and Functional Analysis of Deubiquitinating Enzyme Gene UCHs from Hevea brasiliensis
    Mingyue YUAN, Tianzhong ZHOU, Ma YU, Bin HU, Xiangyu LONG, Hua CHEN
    Bulletin of Botanical Research    2023, 43 (4): 622-630.   DOI: 10.7525/j.issn.1673-5102.2023.04.015
    Abstract1846)   HTML8)    PDF(pc) (1813KB)(1100)       Save

    To investigate the potential function of ubiquitin carboxyl terminal hydrolases(UCHs) in the ubiquitination process of laticiferous of Hevea brasiliensis, the full-length sequences of two UCHs family members(HbUCH-L3 and HbUCH-L5) were isolated from Hevea brasiliensis, and both had typical UCHs domains. The open reading frame of HbUCH-L3 and HbUCH-L5 were 993 bp and 558 bp, and encoded 330 and 185 amino acids, respectively. The results of qRT-PCR showed that HbUCH-L3 and HbUCH-L5 were constitutively expressed in all tissues, but was low in latex. In vitro ubiquitination substrates cleavage of recombinant HbUCHs showed that both HbUCH-L3 and HbUCH-L5 had the function of hydrolyzing ubiquitin. HbUCHs significantly reduced the overall ubiquitination level of C-serum proteins; while, the deubiquitinating activity of HbUCH-L3 was higher than that of HbUCH-L5. Therefore, it was speculated that UCHs played a role in maintaining the dynamic balance of laticiferous ubiquitination and thus played a specific biological function, but the exact mechanism was still unclear.

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    Cloning of WRKY40 Gene from Leymus chinensis and Analysis of Disease Resistance in Transgenic Tobacco
    Danni LI, Jiali LIU, Jitao ZHANG, Baoxiang GU, Fengjin ZHU, Qingjie GUAN
    Bulletin of Botanical Research    2023, 43 (3): 412-420.   DOI: 10.7525/j.issn.1673-5102.2023.03.011
    Abstract3138)   HTML9)    PDF(pc) (3682KB)(100)       Save

    In order to cultivate rice varieties resistant to Xanthomonas oryzae pv. oryzaeRhizoctonia solaniMagnaporthe oryzae and Fusarium fujikuroi, the method of mining resistance genes is an important way to select resistant varieties. RT-PCR method was used to clone the LcWRKY40 gene (MN187915) from the leaves of Leymus chinensis. The result showed that the CDS was 1 053 bp in length and encoded 350 amino acids with a molecular weight of 38.1 kDa. The results of bioinformatics analysis showed that the primary structure of LcWRKY40 contained WRKY domain, zinc finger protein domain and nuclear localization sequence. Phylogenetic tree construction and motif analysis showed that the phylogenetic relationship between LcWRKY40 and HvWRKY38 was closed. The results of subcellular localization in tobacco showed that the LcWRKY40 protein was located in the nucleus, verified by the software prediction. qRT-PCR tissue specific expression analysis showed that LcWRKY40 gene was expressed in root, stem, leaf, leaf sheath, Lemma and anther of Leymus chinensis respectively, but the expression level was the highest in leaf but the lowest in Lemma. Transgenic LcWRKY40 tobacco plants and wild-type tobacco plants were inoculated with X. oryzae pv. Oryzae, R. solani, M. oryzae and F. fujikuroi, respectively, which showed that transgenic LcWRKY40 tobacco plants could alleviate the four pathogens in different degrees, and showed high resistance to M. oryzae and F. fujikuroi. Therefore, it was speculated that LcWRKY40 protein played a key regulatory role in signal pathways such as resisting disease stress and improved the resistance of plant pathogens, which layed a molecular foundation for the study of LcWRKY40 function and abiotic stress.

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    Detailed Phenotypical Analysis on the Mutant ssr1- 2 Encoding a Mitochondrial Protein of Arabidopsis thaliana
    Yuanyuan CAI, Jibenben XIA, Wenhan YING, Jieyao WANG, Tao XIE, Kongya XING, Xuanjun FENG, Xuejun HUA
    Bulletin of Botanical Research    2023, 43 (3): 421-431.   DOI: 10.7525/j.issn.1673-5102.2023.03.012
    Abstract1771)   HTML9)    PDF(pc) (5062KB)(144)       Save

    To explore the role of SSR1 gene encoding a functionally unknown mitochondrial protein of Arabidopsis thaliana in plant growth and stress response, ssr1-2 and its suppressor mutants EMS143 and EMS145 were used to track the growth of their roots and aboveground parts, as well as their proline sensitivity and iron homeostasis, and the effects of ssr1-2 short root phenotype on aboveground growth of seedlings were analyzed by micrografting. The results showed that the primary root length of ssr1-2 was shorter and the root architecture was similar to the fibrous root system. The growth of their shoots was also retarded, but appeared considerably later than the short-root phenotype. The results of micrografting revealed that the roots of ssr1-2 could restrict the growth of the wild-type shoots and, and conversely, the shoot of ssr1-2 could also affect the growth of the wild-type roots, but the former had a bigger effect. ssr1-2 showed a hypersensitive phenotype to proline in seed germination, root length and leaf chlorophyll content. In addition, ssr1-2 appeared insensitive to Fe nutrition, that was, the stimulating effect of Fe salt on seedling growth was significantly less than that of wild-type WS, indicating a significant decrease in its ability to utilize Fe. The results suggested that SSR1 might be involved in the regulation of A. thaliana root growth by affecting the utilization of iron nutrient, implying that the impairment of the mitochondrial iron utilization machinery might contribute significantly to the enhanced inhibitory effect of proline on plant growth and development.

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    The Mechanism of H 2S Signal and Ca 2+ Regulating Stomatal Movement in Medicago sativa
    Xuefeng HAO, Chunxia KANG, Yanxi PEI, Zhuping JIN
    Bulletin of Botanical Research    2023, 43 (2): 281-287.   DOI: 10.7525/j.issn.1673-5102.2023.02.013
    Abstract422)   HTML19)    PDF(pc) (1800KB)(83)       Save

    To explore the role of H2S signal in regulating stomatal movement in alfalfa and the relationship between H2S and Ca2+ during this process, wild type and the calcium transporter mutants of Medicago truncatula were used as experimental materials, and the transcriptional level, cellular level and physiological level were studied respectively, and the expression levels of related genes were compared by qRT-PCR, the content of Ca2+in vivo was detected by fluorescent probe, the content of H2S was measured by electrode method, and the stomatal aperture was observed by microscope respectively. The results showed that compared with the wild type, the content of H2S in the mutant NF3011 and NF2734 decreased significantly; the H2S signal inhibited the expression of calcium transporter encoding gene MTR_6g027580, the exogenous physiological concentration of H2S fumigation could induce stomatal closure of Medicago truncatula, and the combination of treatment with LaCl3, Ca2+ channel blocker, had no effect on stomatal movement of wild type, but opposite in mutant; the concentration of Ca2+ in guard cells was measured by fluorescence probe, which were consistent with the change law of stomatal aperture. In conclusion, the content of Ca2+ in guard cells of leaves is induced by H2S, whose stomatal aperture become smaller at the same time. In this process, the change of intracellular Ca2+ concentration mainly depend on Ca2+ transporter, and small partly depend on Ca2+ channel. The results of this study not only enrich the mechanism of H2S signaling in theory, but also have the potential to be applied to alfalfa production and other crops.

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    Function Analysis of the Transcription Factor PsnbZIP1 of Populus simonii×P. nigra in Response to Salt Stress
    Shixian LIAO, Yuting WANG, Liben DONG, Yongmei GU, Fenglin JIA, Tingbo JIANG, Boru ZHOU
    Bulletin of Botanical Research    2023, 43 (2): 288-299.   DOI: 10.7525/j.issn.1673-5102.2023.02.014
    Abstract302)   HTML22)    PDF(pc) (7255KB)(87)       Save

    To reveal the function of transcription factor PsnbZIP1 from Populus simonii × P. nigra under abiotic stresses, P.simonii × P. nigra was used as material, and the PsnbZIP1 transcription factor gene was cloned with an ORF length of 432 bp, and the molecular mechanism of PsnbZIP1 under salt stress was analyzed. The expression pattern of PsnbZIP1 in P. simonii × P. nigra was analyzed under 150 mmol·L-1 NaCl by q-PCR, it was found that the expression of the gene increased rapidly. The PsnbZIP1 was predicted to be a hydrophilic unstable protein without trans-membrane structure but with signal peptide by bioinformatics analysis. The sub-cellular localization of the gene was observed by Agrobacterium mediated transient expression in tobacco. The results showed that the gene was a nuclear localization protein. It was confirmed by Y2HGold yeast competent cells that the gene had no transcriptional activation activity in yeast. The analysis of the promoter sequence of the PsnbZIP1 gene showed that the promoter contained auxin response elements, ABA response elements, light response elements and seed-specific cis-acting regulatory elements respectively, which might play a role in plant growth and development and response to stress.The promoter also included the MYB binding site and MYBHv1 binding site involved in drought induction, indicating that the gene was likely to interact with some of MYB involved in drought induction.

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    Response Analysis of Transcription Factor PsnbHLH162 Gene in Populus simonii × P. nigra under Salt Stress and Low Temperature Stress
    Senyao LIU, Fenglin JIA, Qing GUO, Gaofeng FAN, Boru ZHOU, Tingbo JIANG
    Bulletin of Botanical Research    2023, 43 (2): 300-310.   DOI: 10.7525/j.issn.1673-5102.2023.02.015
    Abstract472)   HTML17)    PDF(pc) (5711KB)(74)       Save

    To reveal the role of transcription factor PsnbHLH162 of Populus simonii × P. nigra under abiotic stresses, and to explore the signal transduction process of genes in plants, the signal transduction process of this gene in plants is explored to provide the theoretical basis for the acquisition of excellent stress-resistant tree species in the future. In this experiment, PsnbHLH162 gene was cloned and obtained from P. simonii × P. nigra as material, and the target gene and promoter were analyzed by bioinformatics; and the function of genes in response to abiotic stresses was analyzed by fluorescence quantitative PCR. The results showed that the cDNA of PsnbHLH162 gene was 537 bp, and the N-terminal of the gene contained a highly conserved HLH domain, and the encoded protein might be a stable hydrophilic protein without trans-membrane region, localized in the nucleus and had no transcriptional activation activity. The promoter region contained a variety of ABA-responsive, auxin-responsive, light-responsive and circadian elements, confirming that this gene participated in abiotic stress response. The results of fluorescence quantitative PCR showed that under salt stress, the gene expression in root tissue was the highest compared with stem and leaf tissue; under low temperature stress, gene expression in stem tissue was the highest compared with leaf and root tissue. It was found that the expression of PsnbHLH162 could be induced by salt and low temperature in plants.

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    Analysis of the Chloroplast Genome Characteristics of 6 Species of Yucca
    Fei WANG, Wenzhi ZHAO, Zhanghong DONG, Luyao MA, Weiying LI, Zongyan LI, Peiyao XIN
    Bulletin of Botanical Research    2023, 43 (1): 109-119.   DOI: 10.7525/j.issn.1673-5102.2023.01.012
    Abstract3941)   HTML29)    PDF(pc) (5663KB)(6770)       Save

    In order to clarify the characteristics and sequences variation of the chloroplast genome of Yucca, a comparative genomics of the chloroplasts was performed, and a phylogenetic tree based on the chloroplast genome was constructed. The chloroplast genome of Y. treculeana was obtained by high-throughput sequencing technology, and combined with the published chloroplast genome of Yucca, the chloroplast genomes of six species of Yucca were studied by comparative genomics, including basic structure, SSR, boundary contraction and expansion, sequence variation and phylogenetic analysis respectively. The results showed that the chloroplast genome size, gene type and number of the six Yucca species were similar, and the interspecific genome structure was conserved. Several repeated sequences were detected in the chloroplast genome of Yucca, among which SSR loci were mostly composed of mononucleotide, dinucleotide and tetranucleotide, and preferred A and T bases. Three regions with high variation(psbK-psbl-trnS-GCUrpl20-rps12 and ccsA-ndhD) were identified from the chloroplast genomes of six Yucca species according to the nucleic acid polymorphism index(π)≥0.008. The phylogenetic relationships constructed from the chloroplast genome and LSC+SSC region sequences were basically consistent, and the phylogenetic relationships among six Yucca species were confirmed, among which Y. treculeana and Y. queretaroensis were most closely related. The sequencing obtained Y. treculeana chloroplast genome, revealed the genome characteristics and sequence variation of six Yucca species, and clarified the relatedness among various species. The results could provide reference for subsequent molecular marker development and phylogenetic research.

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    Complete Chloroplast Genome Structure and Characterization of Syringa villosa subsp . wolfii
    Yongchang LU, Xin ZHANG, Luyan ZHANG, Jiuli WANG
    Bulletin of Botanical Research    2023, 43 (1): 120-130.   DOI: 10.7525/j.issn.1673-5102.2023.01.013
    Abstract9254)   HTML35)    PDF(pc) (5570KB)(5668)       Save

    To clarify the basic characteristics of the complete chloroplast genome of Syringa villosa subsp. wolfii, the sequence information of chloroplast genome was analyzed by high-throughput sequencing technology, and its systematic evolution position was discussed. The results showed that:(1)The total length of the S. villosa subsp. wolfii chloroplast genome was 156 517 bp, which had a typical tetrad structure and had 131 functional genes, including 36 tRNA genes, 8 rRNA genes and 87 protein-coding genes respectively. (2)The relative synonymous codon usage(RSCU) analysis of the protein-coding region of the chloroplast genome showed that there were 31 codons with RSCU>1, of which 21 ended with A/U base; and there were 34 codons with RSCU<1, including 22 codons ending with G/C base. (3)In the chloroplast genome of S. villosa subsp. wolfii, 334 scattered repeats were detected, including 170 forward repeats and 164 palindromic repeats, and 227 SSR loci were discovered, and PCR primers were designed successfully for 226 of these loci. (4)Phylogenetic tree analysis based on the maximum likelihood method showed that S. villosa subsp. wolfii was the most closely to the S. yunnanensis. In this study, repeated sequences, IR boundaries and phylogeny of chloroplast genome of S. villosa subsp. wolfii were analyzed to provide reference data for molecular markers, phylogenetic classification, species identification and evaluation and DNA barcode development.

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    Cloning and Expression Analysis of MibZIP1 from Macadamia integrifolia
    Haiyun SONG, Tao ZHANG, Peng HE, Shufang ZHENG, Lifeng WANG, Wenlin WANG
    Bulletin of Botanical Research    2023, 43 (1): 131-139.   DOI: 10.7525/j.issn.1673-5102.2023.01.014
    Abstract8272)   HTML28)    PDF(pc) (3902KB)(738)       Save

    In order to clarify the potential functions of bZIP transcription factor family members in Macadamia integrifolia under stress response, MibZIP1 was cloned from ‘Guire 1’ fruit. Combined with the analysis of transcription factor structure and expression patterns under different treatments, it was found that the full length of MibZIP1 was 1 157 bp, and the ORF was 927 bp, and encoded 308 aa,and had a typical bZIP Superfamily domain. Phylogenetic analysis showed that MibZIP1, TsbZIP60 and NnbZIP60 were closely related. The results of tissue expression analysis showed that MibZIP1 expression was the lowest in leaves of ‘Guire 1’ and the highest in branches of ‘695’. The expression of MibZIP1 was significantly up-regulated in ‘Guire 1’ leaves treated by zeatin, salicylic acid, ethephon and abscisic acid respectively, but was unchanged treated by gibberellin and hydrogen peroxide respectively. It was speculated that MibZIP1 was related to the stress tolerance of M. integrifolia, which provided theoretical guidance for the development of new cultivation and hormone regulation techniques.

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    Study on Adaptive Differentiation of Transcription Factor bHLH94 Gene to Altitude in Hippophae neurocarpa and H.tibetana
    Ting QIAN, Fan ZHAO, Yujie ZHANG, Xueli LI, Kun SUN, Hui ZHANG
    Bulletin of Botanical Research    2022, 42 (6): 976-985.   DOI: 10.7525/j.issn.1673-5102.2022.06.007
    Abstract277)   HTML16)    PDF(pc) (5154KB)(250)       Save

    The bHLH proteins are the second large family of transcription factors, which play important roles in plant growth development, physiological metabolism and stress responses. In this study, Hippophae neurocarpa and H.tibetana were used as materials, the positive selection of the transcription factor bHLH94 gene was screened out using transcriptome sequencing. Based on the analysis of gene sequence and expression of HnbHLH94 and HtbHLH94, the response mechanism of bHLH94 gene in H.neurocarpa and H.tibetana adapted to altitude was studied. The HnbHLH94 and HtbHLH94 gene encoded 338 and 335 amino acids respectively. Sanger sequencing method verified the correctness of sequences, however there were 10 non-synonymous mutation sites outside the DNA binding domain of the two genes, which was speculated to be related to the adaptive evolution of the gene. As the altitude increased, the HnbHLH94 gene expression decreased, while the HtbHLH94 gene expression increased based on qRT-PCR technology. It suggested that gene might provide a molecular basis for adaptation to altitude, drought, freezing and radiation respectively, and the sequence structure and expression of HnbHLH94 and HtbHLH94 genes responded to the habitat conditions of elevation.

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    Cloning and Functional Analysis of miR398a from Chrysanthemum× grandiflora in Response to Salt Stress
    Liran YUE, Yingjie LIU, Chenxu LIU, Yunwei ZHOU
    Bulletin of Botanical Research    2022, 42 (6): 986-996.   DOI: 10.7525/j.issn.1673-5102.2022.06.008
    Abstract161)   HTML18)    PDF(pc) (5316KB)(187)       Save

    miRNAs played an important role in abiotic stress, and miR398a and pre-miR398a sequences were obtained from Small RNA database of Chrysanthemum×grandiflora. The sequence alignment showed that the cgr-miR398a was highly conserved with the identified miR398a sequences of other plants. Target genes prediction of cgr-miR398a showed that their annotation of targets include superoxide dismutase[Cu-Zn](CSD2), copper chaperone(CCS), A20/AN1-like zinc finger family protein(SAP8). The qPCR assay showed that the expression levels of miR398a was significantly decreased while target genes showed contrary trend during salt stress in Chrysanthemum×grandiflora. To explore the function of cgr-mir398a in response to salt stress, and an overexpression vector of cgr-mir398a was constructed and transformed into Arabidopsis thaliana.The result showed that overexpression of the cgr-MIR398a in Arabidopsis could decrease seed germination under salt stress and salt tolerance at the adult seedling stage, therefore cgr-miR398a played a negative regulatory role in Arabidopsis in response to salt stress. The results provided a valuable reference for further exploring the functional mechanism of miR398a during salt stress in Chrysanthemum×grandiflora.

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