Previously, we achieved gene editing using CRISPR/Cas9-RNP and plasmid in the PEG-mediated protoplasts transient transformation of rubber tree, and by targeting the HbPDS gene, callus with albino phenotype were obtained, but no edited plants were regenerated because the technology of embryogenesis from callus is not yet mature. In order to obtain gene edited seedlings, we used the same HbPDS target as previous in the callus editing, but used somatic embryos as the transformation receptor instead of callus. After hygromycin resistance screening, 116 positive T₀ generation embryos were selected through Cas9 gene PCR detection, following by next generation sequencing, five embryos were found to be edited at the target, accounting for 4.3% of PCR positive embryos. At last, two regenerated plants were obtained, both were chimeric because only partial albino leaves appeared in the plantlet. Sequencing of both albino and green parts revealed that gene editing had occurred in all samples, besides a homozygous biallelic mutation in one albino leaf, all other leaves exhibited chimeric mutations, with mutant sequences in albino parts accounting for 86% to 100% ratio, while green parts accounting for 66% to 69% ratio. This indicates that the mutation threshold inducing the expected phenotype in rubber tree CRISPR/Cas9 editing plants is higher than 69%, ranging from 70% to 85%, providing theoretical guidance for obtaining gene editing seedlings with expected phenotype in the future. Meanwhile, it is proven that nearly all the regenerated plantlets obtained from T₀ generation somatic embryos are chimeric, thus T₀ generation embryos are not suitable as regenerated materials, but also providing insights to improve the regeneration procedure by using T₁ embryo to get homozygous seedlings in the future. This is the first report about gene editing plants in rubber tree, although they are chimeric, it still enhances the understanding of the function of CRISPR/Cas9 in rubber tree, laying the foundation for improving and applying gene editing technology in rubber tree.

MLO gene is an unique negative regulatory factor for disease resistance in plants, and the mutation in the gene can lead to broad-spectrum disease resistance in plants. In this study, a DNA and cDNA sequence of cassava MLO12 gene were cloned from the entire cassava genome and named MeMLO12. This gene has a total length of 3743 nt and a coding region (ORF) of 1728 nt, with a complete open reading frame containing 15 exons and 14 introns, encoding 586 amino acids. The protein has a molecular weight of 67.2 kDa and an isoelectric point of 8.85. MeMLO12 protein is located on the endoplasmic reticulum membrane, without signal peptides, and forms seven transmembrane domains at 23-45, 74-96, 161-183, 285-307, 312-334, 371-393, 413-435 aa. Quantitative analysis by qRT PCR revealed significant differences in the expression of MeMLO12 gene in resistant and susceptible cassava germplasms after infection with Xanthomonas axonopodis, indicating a negative regulatory effect in the interaction between cassava and Xam. Selecting the 11th exon of the gene for Snap Gene Viewer analysis, 10 455 seed sequences of sgRNA were obtained. Three target sequences of approximately23nt were selected, with a G terminus at the 3′ end of the base composition, and they were constructed onto the CRISPR-Cas9 vector. After verification, it was confirmed that the three target sequences of MeMLO12 have been successfully constructed onto the gene editing vector, named pSGR-Cas9-AT-MeMLO12 vector.

Choline monooxygenase (CMO) is a key enzyme in the synthesis of betaine in higher plants and plays an important role in the physiological process of plant resistance to stress. A CMO gene was identified in banana A and B reference genomes in banana gene database, and the biological information analysis of MaCMO and MbCMO genome sequences showed that MbCMO might be formed by a gene encoding a banana O-fucosyltransferase family protein and MbCMO gene in tandem. The CMO gene coding sequences of Zhanjiang AA (ZJ; AA genotype), Baxijiao (BX; AAA genotype), Guangdong Dajiao (GD; AAB genotype) and Jinfen (JF; ABB genotype) were cloned and compared, it was found that ZJ and BX only contained CMO-A, GD and JF both contained CMO-A, CMO-B1 and CMO-B2, and JF also contained CMO-H. Codon usage characteristics showed that there were 23 common high frequency codons among the four banana CMO gene, and the codons CUU and CCG were the most biased and the weakest codons, respectively. The physicochemical properties of CMO-A protein and CMO-H protein showed that the minimum number of amino acids was 425, the maximum number of amino acids was 470, and the secondary structure of CMO-B1 protein was the most complex. The molecular weight of CMO-B2 protein was 52.02 kDa, and the molecular weight of CMO-A protein was 47.48 kDa. All the four CMO proteins were acidic proteins, which did not have a transmembrane structure and were hydrophilic proteins. Subcellular localization prediction showed that all four CMO were localized in chloroplasts. In terms of evolution, plant CMO had obvious branches in monodicotyledonous plants during evolution, and the CMO-A, CMO-H, CMO-B1 and CMO-B2 proteins of banana were more closely related to other monocotyledonous plant CMO proteins. The results of RT-qPCR showed that CMO expression was up-regulated in the four kinds of banana roots in the early stage of osmotic stress, and the expression levels of ZJ and BX CMO in homozygous A genome were higher than those of GD and JF CMO in heterozygous A and B genomes. CMO expression in the four kinds of banana leaves was down-regulated at the early stage of osmotic stress. The expression levels of homozygous ZJ and BX CMO in A genome were up-regulated at the late stage of osmotic stress, while the expression levels of heterozygous GD and JF CMO in A and B genomes peaked at 10 days and then down-regulated again at 15 days later. It was significantly lower than that of ZJ and BX CMO. This study revealed the differences of CMO genes between the A and B genomes of bananas and the expression patterns of CMO genes in different genotypes of bananas under osmotic stress, which would lay a foundation for further research on the biological functions of CMO genes in the A and B genomes of bananas, especially the relationship between CMO genes derived from different genomes and the ability of banana to resist osmotic stress. It would provide a reference for improving the stress resistance of banana by genetic engineering.

Tigernut (Cyperus esculentus L.), most likely originated from Africa and Mediterranean, represents a novel herbaceous oil crop accumulating high levels of oil in its underground tubers. Water balance is essential for tuber development and metabolism in tigernut. Tonoplast intrinsic proteins (TIPs), which include five phylogenetic groups (i.e. TIP1–5), constitute a subfamily of aquaporin facilitating the fast and passive transport of water across vacuolar membranes. Based on available genome and transcriptome data, one TIP gene named CeTIP2;1 was isolated from tigernut tubers by using the RT-PCR technique. Sequence analysis showed that the full gene length of CeTIP2;1 is 3323 bp, including two introns with a coding sequence of 747 bp; the gene was predicted to encode 248 amino acids with the theoretical molecular weight of 24.73 kDa, the isoelectric point of 5.09, the grand average of hydropathicity of 0.948, the aliphatic index of 114.60, and the instability index of 21.76, implying its stable, acidic, and hydrophobic features, which is consistent with its tonoplast-localization; presence of one conservative MIP domain was observed, which possesses six transmembrane helices, two half helices, and two typical NPA motifs. CeTIP2;1 was shown to exhibit the sequence similarity of 87.20% with AtTIP2;1, which was considerably higher than 46.23% with SoPIP2;1. Further phylogenetic analysis revealed that CeTIP2;1 belongs to the TIP2 group and is a true ortholog of AtTIP2;1. Interestingly, the phylogenetic analysis also supported that TIP2 has diverged into two subgroups sometime before monocot-eudicot divergence. Tissue-specific expression analysis showed that CeTIP2;1 was highly expressed in all tissues examined in this study, i.e., leaf, sheath, root, shoot apex, rhizome, and tuber, with most in tuber/rhizome and lowest in shoot apex. A bell-like expression pattern was observed during tuber development, peaking at the medium stage of swelling and lowest in maturation. Moreover, CeTIP2;1 was also detected in the tuber proteomes, implying its high abundance and key roles. These findings would lay a solid foundation for further uncovering the mechanism of water balance in tigernut tubers.

In order to understand the fruit quality of different varieties of pitaya and to screen the germplasm resources with good quality, 33 pitaya germplasm resources were and 15 botanical traits and quality properties such as single fruit weight, fruit transverse and longitudinal diameters, proteins, total flavonoids, and beet pigments were measured, and fruit quality was comprehensively evaluated by using correlation analysis and principal component analysis. The 33 pitaya germplasm resources had a single fruit mass of 164.67-556.00 g, a transverse fruit diameter of 5.86-9.40 cm, a longitudinal fruit diameter of 7.29-11.94 cm, a fruit shape index of 0.99-1.39, a total sugar content of 78.80-126.05 mg/g, a total phenolic content of 0.64-2.46 mg/g, a total flavonoid content of 0.85-9.67 mg/g, hardness of 6.62-16.86 N, protein content of 0.29-1.45 mg/g, beet pigment content of 0.69-37.01 mg/100 g, and starch content of 14.12-44.60 mg/g. The total acid content ranged from 0.33 to 15.23 mg/100 g. The Pearson’s method was used to correlate the main botanical and quality traits of the pitaya germplasm resources. and quality traits were correlated. The correlation coefficients were 0.478 for total acid and soluble solids, and -0.532 for sugar-acid ratio, while the correlation coefficients were 0.749 and 0.942 for longitudinal and transverse diameters. The correlation coefficient between longitudinal diameter and fruit shape index was 0.426, and the correlation coefficient between weight and sugar-acid ratio and hardness was 0.360 and 0.351, respectively. After principal component analysis, the 15 traits were combined into 7 principal component factors, with a cumulative contribution of 82.55%. The first principal component contributed 22.45%, with single fruit weight, fruit longitudinal and transverse diameter, and sugar-acid ratio contributing more. The second principal component contributed 13.87%, with total sugar content, soluble solids content and starch content contributing more. The contribution of the third principal component was 12.28%, with soluble solids content contributing more. Among them, sugar-acid composition, single fruit weight, fruit transverse diameter, fruit longitudinal diameter, soluble solids content, starch content, amino acid content, etc. are the key indexes for evaluating the comprehensive quality of dragon fruit. After comprehensive evaluation, six excellent pitaya singles with high comprehensive scores were screened, i.e., Reyan 1 hao, Wanhuafenhong, Hongbaolong, Wucihong, Linjiahongyun and Hongjinbao, which can be used as high-quality parents for pitaya quality breeding.

Paphiopedilum is one of the most valuable orchid groups with its unique shape, rich flower colors and long flowering period. The shape of the lip of Paphiopedilum is peculiar, and it is specialized as a pouch or helmet, it looks like a lady’s slippers, so it is also known as the slipper orchid. However, due to the low seed germination rate of Paphiopedilum, low propagation rate of traditional division propagation, immature tissue culture propagation technology, and relatively difficult to cultivate, it is difficult to meet the market demand through artificial breeding. Resulting in the serious exploitation of wild resources and the destruction of its living environment, wild resources continue to decrease, Paphiopedilum is now one of the most endangered species in the world. In this paper, the geographical distribution and habitat of Paphiopedilum in the world and China were analyzed by means of literature retrieval, with special attention to the new species published in recent years, which can provide reference for the conservation and utilization of wild Paphiopedilum germplasm resources. Understanding the geographical distribution characteristics of Paphiopedilum is of great significance for in situ and ex situ conservation of Paphiopedilum germplasm resources. At present, there are about 109 species and 26 varieties of Paphiopedilum in the world. Since the English edition of the Flora of China in 2009, 7 new species of Paphiopedilum have been discovered in China, on the basis of the original 27 species of Paphiopedilum, and 30 new species have been discovered in the world during this period. Paphiopedilum mostly distributed in tropical and subtropical transition areas, originated in southwest China and Southeast Asia, and then spread to Southeast Asian islands, Indonesia (Sulawesi, Java etc.) and China (Yunnan, Guangxi etc.) are the distribution centers of Paphiopedilum. The main factors affecting the distribution of Paphiopedilum are climate, topography, soil, symbiotic microorganisms, pollinators and so on. The distribution pattern of Paphiopedilum is influenced by both large scale (climate factor) and small scale (small habitat, such as landform, soil, etc.). Paphiopedilum may gradually spread and migrate to areas within its temperature tolerance range. In the past 50 years, the high latitude area has warmed more, and the suitable area for the growth of Paphiopedilum has expanded, resulting in its distribution range spreading or migrating to high latitudes and high altitudes. The seasonal temperature changes may cause the distribution of Paphiopedilum plants (Paph. subgenus Brachypetalum) to shift to higher or lower latitudes. The phenology, morphology and genes of Paphiopedilum plants may also undergo certain changes, and the number of species that do not adapt to environmental changes will gradually decrease until extinction, and new species will continue to emerge. The distribution of Paphiopedilum is also affected by topography, soil physical and chemical properties, soil microorganisms and other factors. Microorganisms in soil, especially fungi, can promote the germination of Paphiopedilum seeds. If fungi that are mutually beneficial to Paphiopedilum are lacking in the new habitat, the seeds of Paphiopedilum may not germinate smoothly, and the population of Paphiopedilum may be restricted to a specific area. The distribution of Paphiopedilum is also closely related to pollinators, and the species, quantity and distribution of pollinators affect the distribution pattern of the corresponding Paphiopedilum. Pollinating insects of Paphiopedilum are abundant, mainly belonging to the family of Syrphidae, and insects of Apidae, Halictidae, Colletidae and Megachilidae can also be used as pollinators for Paphiopedilum. There is usually more than one effective pollinator of each Paphiopedilum species. The study on the geographical distribution pattern of Paphiopedilum would provide reliable evidence for the species evolution between genera and within genera. The more primitive Paphiopedilum was mostly terrestrial, while the more evolved Paphiopedilum evolve into semi-epiphytic or epiphytic orchids due to environmental adaptation and competitive pressure, growing on cliffs or between rocks.

As a traditional oleaginous resource on Hainan Island, the camellia species predominantly exist as evergreen trees of small to medium stature. The large-fruit camellia with white flowers is widely distributed across various regions of Hainan Island. This study conducted a survey and sampling of camellia populations aged 30 years and above across all municipalities and counties of Hainan Island. Ploidy identification was carried out using flow cytometry and chromosome squashing techniques. The research revealed that among the 23 populations surveyed, 16 were identified as decaploid and 7 as octoploid, covering 16 municipalities and counties across the island. Decaploid camellia populations were found in 12 counties and cities (Danzhou, Lingshui, Qionghai, Wuzhishan, Ding'an, Chengmai, Tunchang, Lingao, Changjiang, Ledong, Qiongzhong, Baoting), including ancient trees over 100 years old and wild camellia populations in tropical rainforests, all of which were identified as decaploid. In contrast, the octoploid populations were generally under 60 years of age and traced back to artificial cultivation, with no other ploidy camellia resources found in the surveyed populations. It is evident that the camellia resources on Hainan Island are primarily composed of decaploids and octoploids, with the decaploids being indigenous to the island and the octoploids, not exceeding 60 years of age, being introduced from regions such as Guangdong and Vietnam during the 1960s to 1970s. Both types of camellia have demonstrated good adaptability to the growth conditions. Given that decaploid camellia trees have not been found in other camellia distribution areas, the widespread distribution of decaploid camellias on Hainan Island and the discovery of decaploid wild camellia populations in tropical rainforests suggest that the decaploid camellia species are indigenous regional species of Hainan Island, evolved due to unique ecological factors such as the island’s tropical monsoon climate. The distribution range of the resources spans from 18°38′42″N to 19°49′53″N and from 108°51′0″E to 110°29′46″E, at altitudes ranging from 4.17 m to 1090 m.

Through the examination of the phenotypic traits of teak (Tectona grandis L. F.) seeds from diverse provenances and pedigrees, this study elucidated the genetic variability patterns of teak seeds and the mechanisms of response to environmental factor fluctuations, thereby fostering the conservation and genetic improvement of teak germplasm resources. Using seeds from 81 teak pedigrees representing 13 provenances across four provinces (Guangdong, Guangxi, Hainan, and Yunnan) as the experimental material, five traits, namely seed length, width, length-width ratio, length-width product, and hundred-seed weight, were quantified. Significant differences (P<0.01) existed among the five phenotypic traits of teak seeds across provenances and pedigrees, with trait variation coefficients ranging from 10.27% to 23.88% among different pedigrees and from 4.21% to 20.32% among different provenances. Substantial correlations prevailed among various phenotypic traits, with varying degrees of influence on seed traits from different environmental factors. Latitude, altitude, and annual precipitation were the primary determinants affecting teak seed traits, followed by annual average temperature, while the impact of annual average relative humidity was relatively minor. Distinct seed traits also exhibited heterogeneous responses to environmental changes, with seed length, seed width, and length-width product being particularly sensitive to annual precipitation, whereas length-width ratio and hundred-seed weight manifested heightened sensitivity to latitude. Cluster analysis categorized the 13 provenances into three classes. Teak seeds manifest prolific phenotypic variation across provenances and pedigrees, with seed traits being influenced by various factors, including maternal inheritance and geographical climate. The research would provide a theoretical foundation and scientific basis for the conservation and utilization of diverse teak.

Three newly recorded species of China, viz. Annulohypoxylon albidiscum, A. bahnphadengense, and A. spougei, are described and illustrated based on the morphological characters and molecular phylogeny. The specimens were collected from Hainan, China. The detailed morphological features descriptions and photographs were provided for each species. Maximum likelihood analysis and Bayesian inference were used to construct phylogeny trees based on ITS sequences. The three species were supported and confirmed by molecular data. The comparisons between the three Chinese records and holotype specimens were discussed. The specimens are deposited in the Fungarium of Chinese Academy of Tropical Agricultural Sciences (FCATAS).

Soil salinization is an ecological environment issue globally and is one of the major abiotic stress factors that cause crop loss worldwide as well as hindering crop productivity. Halotolerant plant growth-promoting rhizobacteria (PGPR) having potential to promote crop growth and increase salt tolerance in crops can be employed to counter this issue in a more effective and eco-friendly way. The recent study aimed to assess the salinity tolerance and growth-promoting capacity of Bacillus tequilenis Bt-RS isolated from the rhizosphere soil of islands in the South China Sea, for assessment, the corresponding media were used for qualitative and quantitative analysis. A pot trail was also conducted to further evaluated its bacterial consortium to promote banana seedlings growth under salinity stress. The results showed that strain Bt-RS could grow well under the condition of 0-10% NaCl content with various types of salts. The strain Bt-RS also possessed 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity along with indole acetic acid (IAA, up to 6.4 mg/L under the condition of 1%-5% NaCl). Base on the pot experiment, the banana seedlings that were inoculated with Bt-RS displayed a significant increase in height, leaf area, fresh weight of above ground and underground, as well as dry weight of above ground and underground (40.30%, 93.00%, 102.38%, 81.08%, 65.82% and 50.50%, respectively) compared to the control group under normal conditions; The 200 mmol/L NaCl-stress treatment significantly increased the growth of banana seedlings in height, leaf area, fresh and dry weight of both above ground and underground by 20.55%, 65.01%, 98.14%, 144.93%, 87.85% and 54.01% respectively, when compared with the control group. The plant height, leaf area and biomass of banana seedlings were significantly increased in both normal conditions and salt stress environment (P<0.05). There was no difference in plant height and leaf area between the experimental group under salt stress environment and the control group under normal conditions, indicating that applying Bt-RS can alleviated the adverse effects of NaCl stress and enhanced banana seedlings growth attributes. In conclusion, the strain B. tequilensis Bt-RS exhibits high salt tolerance and possess the ability to produce IAA and ACC deaminase, making it a valuable strain for improving banana seedling growth and salt tolerance, and also providing resources of strain for developing and utilizing in saline soil.