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  • Hosted by:Chinese Academy of Sciences
    Sponsored by:Institute of Botany, Chinese Academy of Sciences, Botanical Society of China
    Co-hosted by:Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences
    Institute of Biotechnology and Germplasm Resources, Yunnan AgriculturalAcademy
    Fujian Agriculture and Forestry University
    Hunan Provincial Key Laboratory of Phytohormones and Growth Development, Hunan Agricultural University
    State Key Laboratory of Crops Biology, Shandong Agricultural University

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Current Issue
Editor-in-Chief:Kang Zhong
ISSN 1674-3466 CN 11-5705/Q
Post Code:2-967
Volume 57 Issue 3
01 May 2022
  
INVITED REVIEW
Advances in the Regulation of Rice (Oryza sativa) Grain Shape by Auxin Metabolism, Transport and Signal Transduction
Lixia Jia, Yanhua Qi
Chinese Bulletin of Botany. 2022, 57(3):  263-275.  doi:10.11983/CBB21227
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Rice (Oryza sativa) is a major food crop in the world. The optimization and utilization of the major yield-determining factors are important for increasing yield potential. Among these factors, seed weight is one of the most important factors determining rice production. The heritability of rice grain weight is stable, which is largely unaffected by environmental factors. Grain weight depends on grain shape, which is determined by grain length, grain width, and grain thickness, and the degree of grain filling. The growth of rice glumes and seed endosperm determines the grain shape and weight. The proliferation and expansion of glume cells affect grain development, and endosperm occupies most of the volume of mature seeds. Auxin is an important plant hormone that affects rice yield, which regulates the development of glume and endosperm after fertilization. The spatial-temporal distribution of active auxin is dynamically modulated by auxin metabolism, auxin transport and signal transduction, all of which maintain auxin at the optimal level for seed development. Here we reviewed the research progress of auxin pathways regulating rice grain shape from three aspects, auxin metabolism, auxin transport and auxin signal transduction, to provide clues for exploring the auxin regulation mechanism of grain shape and improve yield in rice.

EXPERIMENTAL COMMUNICATIONS
Cloning and Functional Analysis of Rice Yellow Green Leaf Regulatory Gene YGL18
Kairu Yang, Qiwei Jia, Jiayi Jin, Hanfei Ye, Sheng Wang, Qianyu Chen, Yian Guan, Chenyang Pan, Dedong Xin, Yuan Fang, Yuexing Wang, Yuchun Rao
Chinese Bulletin of Botany. 2022, 57(3):  276-287.  doi:10.11983/CBB22018
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Leaf color mutants are often accompanied by changes in chlorophyll content and abnormal chloroplast structure, and serve as essential materials for studying the functions of chloroplast development and photosynthesis-related genes. In this study, we obtained a yellow-green leaf mutant named yellow-green leaf 18 (ygl18) from Oryza sativa subsp. indica cv. ‘HZ’ with ethyl methanesulfonate (EMS). Compared with the wild type, the leaves of ygl18 turned yellow at three-leaf stage and the degree of yellowing increased as it grew, accompanied by decreasing photosynthetic rate and chlorophyll content. The seed-setting rate, 1 000-grain weight, and effective panicle number were significantly lower than those of the wild type. We observed disordered chloroplast structure, loose stromal lamellas, and stalled development in the mutant using transmission electron microscopy. Genetic analysis indicated that the mutant feature (or phenotype) of ygl18 is controlled by a pair of recessive nuclear alleles, which were located in a 115.2 kb region between markers InDel2 and InDel3 on the long arm of chromosome 3. We found mutations in the 5′UTR of LOC_Os03g48040 encoding FdC2 (ferredoxin C2). The gene’s function on controlling the mutant phenotype was verified using CRISPR transgenic experiments. Our results revealed a genetic basis for leaf color regulatory network and provide new clues for breeding photosynthetically efficient rice varieties in the future.

PAD4 Mutation Accelerating Programmed Cell Death in Arabidopsis thaliana Tyrosine Degradation Deficient Mutant sscd1
Tiantian Zhi, Zhou Zhou, Chengyun Han, Chunmei Ren
Chinese Bulletin of Botany. 2022, 57(3):  288-298.  doi:10.11983/CBB21194
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Programmed cell death (PCD) is extremely important for plant growth and defense. Fumarylacetoacetate hydrolase (FAH) deficient mutant short-day sensitive cell death 1 (sscd1) displayed PCD under short day condition (8 h light/16 h darkness). Our previous study found PCD in sscd1 was related to jasmonates (JAs) signal transduction but not to salicylic acid (SA). Phytoalexin deficient 4 (PAD4) is involved in mutual antagonism between SA and JAs signaling. In this study, PCD was accompanied by up-regulation of PAD4; while mutation of PAD4 accelerated the sscd1 PCD and induced JAs signaling pathway downstream response genes vegetative storage protein 2thionin2.1 and defensin1.2. In triple mutant sscd1/pad4/coil, JAs signal transduction was blocked, resulting in the disappearance of PCD acceleration. PAD4 mutation induce the expression of Tyr degradation gene homogentisate dioxygenase and maleylacetoacetate isomerase, singlet oxygen specific induced genes bonzai1-associated protein 1 and a putative c2h2 zinc finger transcription factor in sscd1, the induction is dependent on JAs signaling receptor COI1. In conclusion, PAD4 mutation increase JAs signaling, then accelerate tyrosine degradation and singlet oxygen accumulation, thereby promoting PCD in the sscd1 mutant.

Changes of Small GTPases Activity During Cucumber Tendril Winding
Binqi Li, Jiahui Yan, Hao Li, Wei Xin, Yunhe Tian, Zhenbiao Yang, Wenxin Tang
Chinese Bulletin of Botany. 2022, 57(3):  299-307.  doi:10.11983/CBB22058
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Tendrils of climbing plants are highly sensitive to mechanical stimuli. Small GTPases signals can be activated by mechanical stimulation which regulate development in Arabidopsis, but whether small GTPases signals play a role in the entanglement of tendrils response to mechanical stimuli is unknown. In this study, the activity of small GTPases in cucumber tendril after mechanical stimulation was investigated using Cucumis sativus, and ROP activity detection results showed that the activity of small GTPases CsROP6 was significantly enhanced during tendril winding. Meanwhile, mass spectrometry revealed that auxin content increased significantly during tendril winding. Further experiments showed that exogenous auxin treatment increased the activity of CsROP6, suggesting mechanical stimulation might activate ROP6 activity through auxin during tendril winding. This study suggests that mechanical stimulation of small GTPases signals may be conserved in different plants.

TECHNIQUES AND METHODS
Establishment and Application of RPA-CRISPR/Cas12a Detection System for Potato Virus Y
Yulong He, Jiage Wang, Shanshan Zhao, Jin Gao, Yingying Chang, Xiting Zhao, Bihua Nie, Qingxiang Yang, Jiangli Zhang, Mingjun Li
Chinese Bulletin of Botany. 2022, 57(3):  308-319.  doi:10.11983/CBB21225
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Plant virus disease is an important factor restricting the safe production of crops. Virus detection can identify viruses and determine the types of viruses, which is the key to disease monitoring, early warning and prevention in crops production. In this study, a detection system based on Recombinase Polymerase Amplification-Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated 12a (RPA-CRISPR/Cas12a) was established for potato virus Y (PVY). The results showed that: (1) Cas12a and other components in the CRISPR/Cas12a detection system were necessary for the detection; (2) The target location of crRNA had a significant effect on Cas12a nuclease activity, and the reaction efficiency was the highest when the target of crRNA contained part of PAM site sequence; (3) The minimum detection limit of RPA-CRISPR/Cas12a was 3×102 copies∙μL-1, which was higher than that of PCR and qPCR methods; (4) The combination of RPA-CRISPR/Cas12a system with crude extraction of nucleic acid and reverse transcription could detect PVY in a non-laboratory setting and the whole process took about 60 minutes. The RPA-CRISPR/Cas12a detection system of PVY established in this study provides an effective method for real-time and rapid visual detection of plant viruses under non-laboratory conditions.

Detection of Reactive Oxygen Species Using H2DCFDA Probe in Plant
Haitao Hu, Tingting Qian, Ling Yang
Chinese Bulletin of Botany. 2022, 57(3):  320-326.  doi:10.11983/CBB22043
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Reactive oxygen species (ROS) are a ‘double-edged sword’ in plants. On the one hand, ROS, as a signal molecule, plays pivotal roles in many aspects of life activities; on the other hand, excessive accumulation of ROS can cause oxidative damage to biological macromolecules. Accurate detection of ROS is essential to assess its intracellular redox status. Due to the characteristics of short half-life and strong reactivity of ROS components, their qualitative and quantitative analysis are difficult. It is critical to select the appropriate detection method and improve the spatiotemporal accuracy of detection for research in plant sciences and in other fields. At present, fluorescent probe analysis has attracted the attention of researchers because of its advantages of high sensitivity, good selectivity, low detection limit and strong intuition. This article introduces the detailed operation protocol and attentions for ROS detection using 2′,7′-dichlorodi-hydrofluorescein diacetate (H2DCFDA) fluorescent probe based on flow cytometry and confocal microscope. These methods can be used to detect ROS levels and distribution in model plant tissues, including Oryza sativa, Arabidopsis thaliana, Zea mays and Glycine max.

Establishment and Application of a Rapid Genetic Transformation Method for Peanut
Qiong Zhai, Rongqin Chen, Xiaohua Liang, Chuchun Zeng, Bo Hu, Ling Li, Xiaoyun Li
Chinese Bulletin of Botany. 2022, 57(3):  327-339.  doi:10.11983/CBB21192
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Plant transformation is an important tool for genetic engineering. The key technology of genetic transformation is to introduce foreign genes into plants genomes quickly and efficiently and reduce the time to obtain transgenic offspring. Peanut (Arachis hypogaea) is an important oil and cash crop in China. The genetic transformation system is still not well established in peanuts. It seriously restricts the function research of peanut genes and the molecular breeding progress. Here, we established a rapid and stable genetic transformation system in peanuts. The Agrobacterium tumefaciens was injected into the second stem of peanut to produce transgenic plants. Then positive transgenic peanut was transplanted and tamped backfill to cover the injection point. Those seeds above the injection point were picked for further screening and analyzing. The results showed that over 40% of transgenic plants were obtained and displayed chimeric in T0 generation. The T0 seeds were harvested about 5 months after rapid-transformation. About 9% of T1 peanuts were hybrids rather than chimeras. To solve the problem of few seed in some transgenic plants, the rapid-transformation system was combined with traditional tissue culture. This rapid-transformation system has potential value in garlic (Allium sativum), potato (Solanum tuberosum), and freesia (Freesia refracta). Altogether, this study establishes a rapid and stable genetic transformation system for peanuts, which sheds light on other plants’ genetic transformation.

An Efficient Protoplast Transient Expression System in Camellia sinensis var. sinensis cv. ‘Tieguanyin’
Yuqin Zhang, Jiacheng Wu, Meng He, Renyi Liu, Xiaoyue Zhu
Chinese Bulletin of Botany. 2022, 57(3):  340-349.  doi:10.11983/CBB21206
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Recent advances in the genomic sequencing of tea plants have laid the foundation for tea research at the molecular and gene levels. However, the transgenic technologies are immature and the life cycles are long for tea trees, it is still difficult to conduct functional analyses of tea genes. This study used young leaves of Camellia sinensis var. sinensis cv. ‘Tieguanyin’, established a useful formula by testing multiple concentration combinations of cellulase, pectinase, macerozyme and mannitol. By evaluating the quantity, viability and debris of resulted protoplast, we successfully established a highly efficient mesophyll protoplast isolation and PEG-mediated transient expression system in Tieguanyin seedling leaves, with a transformation rate reaching 56.25%. Using this system, the subcellular localization of two pivotal enzymes in the theanine metabolism pathway (the theanine synthetase (TSI) and the glutamine synthetase (GSII-1.1)) were explored. Results show that, these two enzymes are both localized in the cytosol of the Tieguanyin protoplasts. Together, the establishment of this tea mesophyll protoplast extraction and transient expression system would lay a technological foundation for studying the function of tea genome.

Original article
Rapid Propagation Technology and Field Production of Hemerocallis fulva cv. ‘Fenmeiren’
Lü Xiuli, Yu Zequn, Chen Xiangbo, Fu Renjie, Miao Shanshan, Du An
Chinese Bulletin of Botany. 2022, 57(3):  350-357.  doi:10.11983/CBB21182
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Using the scape as explants, we successfully established the tissue culture and rapid propagation technology for the large-scale production of Hemerocallis fulva cv. ‘Fenmeiren’. Our results showed that the survival rate of explant was 95%, which explants were obtained in June and sterilized in the 15% (v/v) sodium hypochlorite solution for 8 minutes. The best proliferation medium was MS + 1.0 mg·L-1 6-BA + 0.004 mg·L-1 TDZ + 0.1 mg·L-1 NAA. After 30 days of cultivation, the monthly proliferation coefficient reached 2.9. On the medium of MS + 0.1 mg·L-1 6-BA + 0.1 mg·L-1 IBA, the tissue culture seedlings did not differentiate and grew vigorously. The optimal rooting medium was 1/2MS + 0.4 mg·L-1 IBA + 20 g·L-1 sucrose, and the rooting rate was 95%. by use of perlite:peat=1:2 (v/v) as the transplanting matrix, the survival rate of transplanting was 85% and the qualified rate was 75%. At present, the large-scale production of this cultivar has been achieved and 2.0 × 105 tissue culture seedlings were produced, which all performed well in the field.

SPECIAL TOPICS
Plant Kinesin: from Microtubule Arrays to Physiological Regulation
Yunhui Wang, Yifan Wang, Jiayu Lin, Jinhong Li, Shien Yao, Xiangchi Feng, Zhenlin Cao, Jun Wang, Meina Li
Chinese Bulletin of Botany. 2022, 57(3):  358-374.  doi:10.11983/CBB22007
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Kinesins are molecular motors that move along microtubules tracks, catalyze the hydrolysis of ATP to ADP, convert the chemical energy stored in ATP molecules into a mechanical force efficiently, and play important roles in various life activities such as cell morphogenesis, cell division, cell movement, intracellular transport, and signal transduction. Plant kinesin research lag behind that of animals and fungi, not only because plants have evolved a unique kinesin family, but also have more members than that of animals. Here we summarize the most recent advancements made towards understanding kinesin functions in the dynamic organization of microtubule arrays, including cortical microtubules and mitotic pre-prophase band, spindle apparatus and phragmoplast, and the regulation of plant physiological activities. We also performed a systematic analysis, classification and functional prediction of kinesins in soybean, the important cash crop, and found that the numbers of soybean kinesins are largely expanded. Taking advantage of the soybean transcriptome data in the public database, the functions of some soybean kinesins were predicted, to provide some clues for the study of kinesin functions in soybean and other crops.

Coupling Regulation of Endocytosis and Exocytosis in Plants
Xu Yan, Mei Xu, Yutong Wang, Weihuai Pan, Jianwei Pan, Jianxin Shou, Chao Wang
Chinese Bulletin of Botany. 2022, 57(3):  375-387.  doi:10.11983/CBB21223
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Eukaryotic cells transport macromolecules and particulate matter away or to the plasma membrane through endocytosis and exocytosis, including certain proteins with important biological functions. The coupling between these two vesicle transport pathways is essential for maintaining the integrity of the plasma membrane as well as regulating the abundance and activity of plasma membrane proteins. In animals, the spatiotemporal coupling of synaptic vesicle endocytosis and exocytosis has been shown to be necessary for the continuation of neurotransmission. In recent years, increasing evidences on the plant vesicle trafficking show the existence of a coupling regulation between endocytosis and exocytosis, which plays an important role in plant growth and development as well as responses to the environment. Here we summarize the physiological significance of plant cooperative regulation of endocytosis and exocytosis, and discuss the potential coupling mechanisms based on the recent progress in the study of clathrin-mediated vesicle trafficking.

The Structure and Function of Leaf Veins and Their Influence on Leaf Economic Spectrum
Yiling Wu, Fanglan Li, Hui Hu
Chinese Bulletin of Botany. 2022, 57(3):  388-398.  doi:10.11983/CBB21203
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Vascular tissues inside the mesophyll and peripheral mechanical tissues constitute the veins. The diverse orders and network structures of veins contribute to their functional diversification and differentiation. In this review, we summarized the research progresses on the structure and function of the leaf vein system. We reviewed three aspects of veins and critically evaluated the characteristics of the leaf vein hierarchy system and its important role in leaf economic spectrum (LES), and explained the mechanisms linking vein traits and other functional traits of the leaf. Leaf veins of different orders show obvious functional differentiation in terms of hydraulic conduction and mechanical support. Among them, the first three orders of veins (major veins) play a major role in maintaining leaf shape, leaf surface area and physical support, and which is conductive to the growth of leaves with the largest light-receiving area. The higher order veins (minor veins) have the function of water regulation, and their coordination with the stomata determines the rate of leaf water transport, transpiration and photosynthesis. The patterns of dynamic variation in leaf spread and leaf vein development explain the relationship between vein density and leaf size. Leaf surface area is negatively correlated with the density of main veins and positively correlated with the diameter of main veins, but independent of the density of minor veins. The framework model of LES linking with vein traits predicts that leaves with higher vein density have short lifespan and smaller leaf mass per area, which explains the better leaf carbon assimilation rate, metabolism rate and resource acquisition strategy with higher leaf vein density.

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