MNP (multiple nucleotide polymorphism, MNP) is a new molecular marker technology emerged with the development of molecular marker technology. While genotyping by target sequencing (GBTS) amplifies only one SNP site in one amplicon, MNP can amplify multiple SNP sites simultaneously in one amplicon based on GBTS. This technology has the features of low cost, high detection efficiency, flexible application and wide adaptability, Compared with SNP, MNP has better ability to differentiate varieties. In order to establish a rapid, accurate and efficient method for cassava variety identification and to screen MNP molecular markers suitable for cassava variety identification, this study screened highly polymorphic regions and designed primers in the whole genome range, and finally obtained 623 cassava MNP marker loci in the whole genome range, the cassava MNP markers were evaluated using 28 cassava varieties. The results showed that the MNP marker typing reproducibility was up to 100%. 4.07±1.68 alleles were detected in 28 cassava varieties, with a maximum of 12 alleles. When all cassava varieties were compared one by a one, 99.47%(376/378) of the differences between pairs were greater than 46%, with the proportion ranging from 0.3% to 81.0%, with a mean value of 71.78%. Meanwhile, MNP markers can be used for identification in the breeding process. In conclusion, the cassava MNP molecular markers developed in this study have high reproducibility, polymorphism and variety differentiation ability, and could be widely used for research on germplasm diversity, new variety breeding and variety identification of cassava.

The molecular mechanism of the formation of flower diversity is still unclear. In this study, the transcriptome and microRNA sequencing of Dendrobium officinale and D. loddigesii flower organs were performed, and the miRNA-mRNA network and target protein interaction network were constructed by cytoscape and compared at the molecular level. The results of transcriptome studies showed that mRNAs related to flower development had MADS-box AP3 gene, hormone genes, etc. The miRNA-mRNA network analysis showed that the differentially expressed miRNAs and mRNAs were involved in the differential formation of floral pattern, and miR5179 and its target genes AP3, miR160 and their target genes ARFs, miR164 and its target genes NAC021, miR159 and the target genes GAM1, bdi-miR159 and the target genes CKX9, miR167b-3p and the target genes ANT may be involved in floral pattern formation, and the above miRNA was upregulated in D. officinale. The protein interaction pathway showed that two kinds of dendrobium had the key gene of AP3, D. loddigesii lacked four genes: CUL1, TIR1, TIR1A and RBS; there weret wo differentially expressed ARFs genes in the auxin response interaction pathway of two Dendrobium, and D. loddigesii lacked one ARFI gene; D. loddigesii flora specifically possessed the protein interaction pathway of auxin signal transduction target genes constituted by the NAC021 gene. Differential expression of miRNAs/mRNAs in D. officinale and D. loddigesii and protein interaction of target genes may be the main mechanisms leading to the difference between the two Dendrobium flower types.

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.

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.

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.

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.

Mutagenesis breeding is a fast and effective breeding method, which has special important significance for the improvement of ornamental plant varieties. Orchids have important ornamental, medicinal, edible and/or cultural value, and broad market prospects. Studies have shown that mutagenesis causes the character changes including plant type, leaf character, flower number, flower size, flower type, flower color, flowering time, ornamental period, insect resistance, disease resistance, stress resistance in orchids, and to date, at least 930 mutants and 16 new orchid cultivars have been generated. This paper reviews the achievements of mutagenesis breeding research in orchids, clarifies current situation and factors affecting the effect of orchid mutation breeding, summaries mutation mechanism of orchids, and finds the methods to further improve the efficiency and effect of orchid mutation breeding. It would provide references for creating new varieties of orchids by better utilizing mutation breeding technology and further clarifying the mutation mechanism of orchids.

Paphiopedilum hirsutissimum (Lindl. ex Hook. f.) Stein is a rare and endangered plant in China with high ornamental value. In order to explore the genetic characteristics of wild P. hirsutissimum resources in Southwest China, and contribute to the protection and utilization of the wild resources, this study used SSR molecular marker to analyze the genetic diversity and population genetic structure of 190 P. hirsutissimum resources which collected from six populations in Guangxi, Yunnan and Guizhou provinces in Southwest China. In this study, the results showed that ten pairs of primers with good amplification effect were selected from 115 pairs of primers, and 50 alleles were detected by 10 pairs of SSR primers of 190 P. hirsutissimum. The average number of alleles (N_a) was five and the average number of effective alleles (N_e) was 2.4835. The average Shannon information index (I) was 0.8592. The average observed heterozygosity (H_o) was 0.4518 and the average expected heterozygosity (H_e) was 0.4387. The average polymorphic information content (PIC) was 0.3996 and the average Nei's expected heterozygosty (h) was 0.4370. In this six wild P. hirsutissimum populations, the number of alleles (N_a) was from 2.8000 to 4.3000. The number of effective alleles (N_e) was from 1.9655 to 2.4060. The observed heterozygosity (H_o) was from 0.3891 to 0.4839. The expected heterozygosity (H_e) was from 0.3795 to 0.4683. The Shannon information index (I) was from 0.6584 to 0.8369, and the Nei's expected heterozygosty (h) was from 0.3648 to 0.4382. In the six wild populations, the genetic diversity of Guangxi Yachang (GYC) and Guizhou Wanfeng Lake (QWF) populations was generally higher (h=0.4382, h=0.4276), while the genetic diversity of Guangxi Mulun (GML) population was relatively low (h=0.3648). Analysis of molecular variance (AMOVA) results showed that genetic variation mainly occurred among individuals within the population (94%), while genetic differentiation within populations was very small (6%). UPGMA cluster analysis based on genetic distance showed that the genetic distance of the six P. hirsutissimum populations was very small, there was no obvious group division and the genetic distance was not completely related to the geographical location. The results of Structure and principal coordinate analysis were consistent with those of UPGMA cluster analysis. Structure and principal coordinate analysis results showed that there was homogenization among the populations, and there was no obvious group division. In summary, the genetic diversity of Paphiopedilum hirsutissimum resources is relatively rich, which can provide a theoretical basis for the protection and utilization of wild P. hirsutissimum resources in Southwest China.

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.