In the Orchidaceae family, Paphiopedilum exhibits extraordinary ornamental value and promising market prospects due to its unique appearance. However, Paphiopedilum is at risk of extinction worldwide due to slow growth rate, high environmental demands, and continuous collection by humans. Artificial seedlings can serve as a potential solution for ex situ conservation, reintroduction and commercial production. Unfortunately, artificial propagation is challenged due to low seed germination rate, slow growth rate, and limited resistance to ecological environments. Mycorrhizal symbiosis between plants and fungi has played a crucial role in the natural development of Paphiopedilum, particularly endophytic fungi, which significantly contribute to seed germination and plant growth. This article presents an analysis of features of endophytic fungi of Paphiopedilum, methods of endophytic fungi separation, types and functions of research, and the application prospects, while also outlining future research direction for the conservation of wild Paphiopedilum, seed germination and commercialization. This study aims to provide a comprehensive reference for future research on Paphiopedilum conservation, seed germination and commercialization.

NAC transcription factors are a large gene family in plants which play important roles in regulating plant growth and development, signal transduction, stress response, etc. In the previous study, the NAC gene, named MiNAC7, was isolated by the yeast two-hybrid system. In this study, the bioinformatics, expression patterns and gene functions of genes were studied. Bioinformatics analysis showed that the MiNAC7 gene was located on chromosome 10, with 4 introns and 5 exons. The length of the coding region of the MiNAC7 gene was 1137 bp, encoding 379 amino acids, the theoretical isoelectric point was 4.88, the molecular weight of the protein was 93.41 kDa, and the amino acid sequence contained a conserved NAM domain. Phylogenetic tree analysis showed that mango MiNAC7 and pistachio PvNAC26 had the closest genetic relationship and the highest homology, and the amino acid sequence similarity was 69.64%. Promoter sequence analysis showed that the promoter region of the MiNAC7 gene contained light response elements, gibberellin response elements and auxin response elements. Expression analysis showed that the expression level of the MiNAC7 gene was high in the stems and buds of juvenile tissues, low in flowers, high in the stems of adult tissues, and low in flowers and leaves. At the same time, it was found that the MiNAC7 gene maintained a high expression level in the leaves at the vegetative growth stage and a low expression level in the leaves at the flowering transformation stage and flower development stage. Overexpression of the MiNAC7 gene led to a late-flowering phenotype in transgenic Arabidopsis. The overexpression of the MiNAC7 gene in Arabidopsis significantly reduced the expression level of the floral-promoting genes AtFT and AtAP1, while the expression level of the late-flowering gene AtFLC was significantly increased. Stress treatment showed that Arabidopsis with excessive expression of the MiNAC7 gene improved its resistance to drought and salt and improved its resistance to GA₃ but was more sensitive to ABA. This study shows that the mango MiNAC7 gene not only affects flowering but also participates in the response to abiotic stress, which would lay a foundation for further research on the gene regulatory network of the MiNAC7 gene involved in regulating mango flowering and stress response.

Black pepper (Piper nigrum L.) is a world-famous spice crop, known as ‘the king of spice’. The establishment of tissue culture and regeneration technology of black pepper has important application value in improving the efficiency of seedling breeding and industrial development. Here, we investigated the effects of explant sterilization, callus induction, and embryo differentiation in somatic cell regeneration systematically in black pepper, and established the tissue culture technology using the elite cultivar ‘Reyin-1’ as the material. The results showed that episperm micropylar tissues could be employed as the explants. The ripe fruits were first sterilized with 75% alcohol for 45 s, and then peeled seeds were sterilized for 45 s in 75% alcohol, followed by 6-10 min of soaking in 0.1% mercuric chloride. The episperm was removed from the plantlet after two weeks culture in darkness, which was used as the explant for somatic embryogenesis. The callus was induced from episperm micropylar tissues within two-month subculture. The optimal culture medium for callus induction and embryogenic callus differentiation was MS+1.5% sucrose+0.80 mg/L 2,4-D+1.200 mg/L KT, and the callus induction rate was 88.14 %, and differentiation rate of embryogenic callus was about 58.18%. The embryo callus were transplanted on somatic embryo induction medium. Somatic embryos were induced on the medium MS+1.5% sucrose+0.40 mg/L 2,4-D+0.600 mg/L KT, with a induction rate of 13.33%. After two months of rooted culture, tissue culture plantlets were obtained, and the optimal medium for rooting culture was 1/2MS+1.5% sucrose+0.25% activated carbon. In conclusion, the study provided a combine method of alcohol and mercuric chloride for explants sterilization, identified that micropylar tissues of the episperm was the best explants for somatic embryogenesis in black pepper. The medium and culture conditions of callus induction, embryo differentiation, rooting culture during somatic embryogenesis were further explored in black pepper. The results would lay a foundation for high-efficiency tissue culture seedling and transgenic breeding in black pepper.

Six nuclear extracts and three leaves of different development stage were screened to establish the flow cytometry (FCM) method for determining the genomic size of Piper L. and Cucumis sativus L. was used as the DNA reference standard. Result showed that the modified nuclear extract and inverted leaf were suitable for the flow cytometry of Piper L. The genome size of cultivated varieties, including Kuching, Aman and EMAS, ranged from 667.94 to 719.32 Mb, and Reyin1, microphylla variety from Cambodia, Panniyur-1 and 73F5 ranged from 759.69 to 785.38 Mb. Therefore, the genome size was significantly different among cultivated varieties. Furthermore, it is speculated that the chromosome numbers of P. wallichii and P. hancei, with genome size larger than 900 Mb, should be larger than 52 with a multiple of 13, and the number of chromosomes of germplasm, with genome size ranged from 600 to 800 Mb, may be 52. The hybrid not only had similar phenotypes to cultivated varieties and P. flaviflorum, but also had an average genome size of cultivated varieties and P. flaviflorum. Therefore, it is speculated that the hybrid may be the hybrid offspring of cultivated varieties and P. flaviflorum. This study would provide a method for the determination of the genome size of Piper L. germplasm, and provide a technical basis for the study of Piper L. germplasm evaluation and distant hybridization.

In order to understand the diversity and distribution characteristics of orchids in Gexigou National Nature Reserve in Sichuan province, a systematic survey was conducted using the line transect combined with sample sampling method from 2020 to 2021. A total length of 95 km including 17 lines were set up, covering various function, habitat and elevation of the reserve. 34 species of Orchidaceae in 20 genera were investigated. Among them, 10 were rare and endangered species, five were wild species under second-class national protection; and two (Liparis brunnea and L. rostrate) were new record species in Sichuan province. Most of them were mainly in terrestrial orchid of life type and absolutely dominant in north temperate zone of geographical. A total of 6703 orchids of 34 species were investigated and distributed in 290 sites. Galearis spathulata and Neottia megalochila were the most and least abundant species respectively, and Epipactis helleborine was the most widely-distributed. Among them, the height of Cypripedium guttatum showed a significant negative correlation with elevation, while there was a slightly correlation in other species. Most of Orchidaceae genera were distributed in 3100-3300 m, followed by 2900-3100 m, while the maximum number of individuals were distributed in 3900-4100 m. The number of Cephalanthera longifolia was positively correlated with slope significantly, the number of Galearis spathulata was positively correlated with elevation significantly, the number of Herminium lanceum was positively correlated with vegetation significantly, the number of whole genus of Herminium was positively correlated with elevation moderately. In general, the orchid resources in Gexigou Nature Reserve are relatively rich, and more distribute in the middle elevation. The number of some species is significantly correlated with environmental factors. Compared with the previous investigations in this reserve, the number of orchid species is significantly increased. However, three national protected species of Cypripedium with high ornamental value were not found, and the population of one species of Cypripedium decreased. Therefore, it is urgent to protect, publicize and enforce the law on orchids in the reserve.

The study aimed to define the floral colour of Phalaenopsis precisely and establish a classification system of Phalaenopsis based on floral colour phenotypes. In this study, the floral colour phenotypes of 146 Phalaenopsis germplasm resources were determined using colourimeter and RHSCC, and the floral colours of each Phalaenopsis were named by cluster analysis and combined with ISCC-NBS system, and the quantitative classification was studied. The results showed that the colour of Phalaenopsis petals and sepals did not differ much, and the lip petals were darker and more colourful than petals and sepals. The classification results based on cluster analysis of L^*, a^* and b^* of petals could not fully characterize the classification of Phalaenopsis flower colours. The ISCC-NBS system divided Phalaenopsis flower colour into yellow, brown, red, violet, pink, purple and white groups. Each group had a preferable correspondence with L^*, a^* and b^* parameters of the CIE Lab system, which could realize the quantitative description of different Phalaenopsis flower colour and the colour group classification was reasonable. Phalaenopsis were rich and colourful, and there were significant differences between different colour groups of Phalaenopsis, but lacking blue-green colour. Overall the brightness and colouration of Phalaenopsis were negatively correlated, and could be divided into two groups, the first type containing yellow, brown, violet, pink, purple and white groups, the second type containing red colour groups.

Phalaenopsis I-Hsin Venus is an excellent variety of fragrant flowers, with elegant flower types, long flowering period and rich floral fragrance, which has high garden ornamental value. ACT1, ACT2, ACT3, GAPDH, EF1α, TUA, TUB and Ubi, were selected as the candidate internal reference genes based on the transcriptome data of different flower development stages in Phal. I-Hsin Venus to select the appropriate reference genes for RT-qPCR analysis of the correlated genes in the biosynthesis pathway of the floral scent in Phalaenopsis I-Hsin Venus. The expression of the candidate internal reference genes in inflorescences at different stages was detected by RT-qPCR. The candidate internal parameter genes were analyzed by combining three internal parameter gene stability analysis software: geNorm, NormFinder and BestKeeper. The results of comprehensive analysis showed that ACT1 was the most stable and could be used as the best internal reference gene for the expression analysis of Phal. I-Hsin Venus.

In order to further investigate the anthocyanin biosynthetic pathway in Phalaenopsis-type Dendrobium (Den-Phals) varieties with different flower color, this study used widely targeted metabolomics technology to analyze the anthocyanin composition and content of five Den-Phals varieties with blue-purple, light peach-red, red, purple-red and deep purple-red floral color, respectively. The results showed that a total of 38 metabolites were identified from the tested Den-Phals, most of which were glycosides and acylated derivatives of cyanidin (Cy), peonidin (Pn), pelargonidin (Pg), delphinidin (Dp) and petunidin (Pt). There were significant differences in the composition and content of anthocyanin in five Den-Phals varieties. Metabolite difference analysis showed that Cy and Dp glycosides in purple flowers were significantly higher than those in light peach red flowers. The contents of three Dp-type glycosides in blue-purple flowers were significantly higher than those in other colored varieties. Only one Cy-type glycoside was screened out, and the content of which in light pink flowers was high. The content of Cy-type glycosides was the highest in most samples. With the increase of cyanidin and delphinidin and the derivatives, the color of flowers deepened and turned to red-purple. It was speculated that Cy-type glycosides made Den-Phals tend to purple while Dp glycosides gave Den-Phals a blue-purple tone. Based on the above results, it is speculated that there are acylation synthesis pathways of the above five anthocyanins in Den-Phals, which can provide a basis for flower color formation mechanism and flower color improvement of Den-Phals.

The specific transport routes of photosynthetic compounds affect the resource allocation of nutrients at different growth stages. The most important is to affect the quality of flowering and fruit, and promote the artificial cultivation and production of ornamental plants. In this study, Pleione formosana, an endangered species, was used as the material, carboxylfluorescein (CFDA) tracer and laser scanning confocal microscopy were used to study the assimilate transport directions in four different growth and development periods, namely, dormancy period, blossom period, vigorous growth period and half dry period to understand the function of source and reservoir of different organs in different periods. During the dormancy period, assimilates were introduced from mother pseudobulb to bulb plate and finally transported to bud for the growth and development of new bud. In the blossom period, the assimilates were transported irreversibly from the mother bulb to the flower organs to ensure the quantity and quality of flowering. In the vigorous growth period, the assimilates mainly supplied the growth of the leaf and new bulbils at the apex of the mother pseudobulb. In the half dry period, the two daughter pseudobulbs competed with each other for nutrients from the mother pseudobulb, while some assimilates were still used to satisfy the growth and development of apex bulbils. The results showed that pseudobulb, as a nutrient storage organ, played an important role in assimilate distribution and generation regeneration succession in four different periods. Its special function would provide reference for artificial cultivation and commercial production in the future.

Ploidy identification is one of the basic tasks for germplasm resources and new variety breeding of tropical flowers. In this study, flow cytometry was used to identify the ploidy level of Cymbidium hybrid seedlings. aiming at the key links such as the type of dissociated liquid, the tissue organ of the test material, evaluation indexes such as the nuclear dissociation, the clear degree of the main peak, the easy degree of sampling, and the coefficient of variation. A technique system was established and the stomatal traits of different ploidy were analyzed. The results showed that CyStain UV Precise P was the best of the three dissociated liquid and young leaves were most suitable. Six triploid and two tetraploid were identified from the offspring of the three hybrid combinations. There were significant differences in stomatal length, width and density between different ploidy (P<0.05). The rank of stomatal length and stomatal width was tetraploid>triploid>diploid, while stomatal density was reversed. This study is valuable for the polyploid selection and use of different ploidy germplasm resources for cross breeding of Cymbidium hybrid.