Mei (Prunus mume) is widely cultivated in eastern Asia owing to its favored ornamental characteristics and its tolerance for low temperatures. Reverse transcription quantitative real-time polymerase chain reaction (qRT-PCR) is a widely used method for gene expression analysis, requiring carefully selected reference genes to ensure data reliability. The aim of this study was to identify and evaluate reference genes for qRT-PCR in mei. Ten candidate reference genes were chosen, and their expression levels were assessed by qRT-PCR in four sample sets: 1) flowering mei; 2) mei undergoing abiotic stress; 3) different genotypes of Prunus species; and 4) all mei samples. The stability and suitability of the candidate reference genes were validated using commercially available software. We found that protein phosphatase 2A-1 (PP2A-1) and PP2A-2 were suitable reference genes for flowering with ubiquitin-conjugating enzyme E2 (UBC) also being suitable for different genotypes of Prunus species. UBC and actin (ACT) were most stably expressed under abiotic stress. Finally, the expression of an AGAMOUS homolog of Arabidopsis thaliana (PmAG) and a putative homolog of Group 2 late embryogenesis abundant protein gene in A. thaliana (PmLEA) were assessed to allow comparisons between selected candidate reference genes, highlighting the importance of careful reference gene selection.
Tao Wang, Ruijie Hao, Huitang Pan, Tangren Cheng and Qixiang Zhang
Yushu Li, Zongda Xu, Weiru Yang, Tangren Cheng, Jia Wang and Qixiang Zhang
The MADS-box gene SOC1/TM3 (suppressor of overexpression of constans 1/tomato MADS-box gene 3) integrates multiple flowering signals to regulate the transition from vegetative to reproductive development in arabidopsis (Arabidopsis thaliana). Although SOC1-like genes have been isolated from a wide range of plant species, their orthologs are not well characterized in mei (Prunus mume), an important ornamental and fruit plant in east Asia. To better understand the molecular regulation of flower development in mei, we isolated and characterized three putative orthologs of arabidopsis SOC1, including PmSOC1-1, PmSOC1-2, and PmSOC1-3. The phylogenetic tree revealed that these genes fall into different subgroups within the SOC1-like gene group, suggesting distinct functions. PmSOC1-1 and PmSOC1-3 were mainly expressed in vegetative organs and at low expression levels in floral parts of the plants, whereas PmSOC1-2 was expressed only in vegetative organs. Furthermore, the expression level decreased significantly during flower bud differentiation development, suggesting a role for these genes in the transition from the vegetative to the reproductive phase. Overexpression of PmSOC1-1, PmSOC1-2, and PmSOC1-3 in arabidopsis caused early flowering. Early flowering also increased expression levels of four other flowering promoters, agamous-like 24 (AGL24), leafy (LFY), apetala 1 (AP1), and fruitfull (FUL). Moreover, the overexpression of PmSOC1-1 and PmSOC1-2 resulted in a range of floral phenotype changes such as sepals into leaf-like structures, petal color into green, and petal into filament-like structures. These results suggested that the genes PmSOC1-1, PmSOC1-2, and PmSOC1-3 play an evolutionarily conserved role in promoting flowering in mei, and may have distinct roles during flower development. Our findings will help elucidate the molecular mechanisms involved in the transition from vegetative to reproductive development in mei.
Le Luo, Yichi Zhang, Yingnan Wang, Tangren Cheng, Huitang Pan, Jia Wang and Qixiang Zhang
Gesnariad (Primulina yungfuensis) is a popular houseplant species, native to southwest China. However, stunting frequently occurs as a result of limited knowledge about the growth requirements of this plant. Understanding water and fertilizer requirements of gesnariad are important for effective large-scale greenhouse cultivation. Using a response surface methodology (RSM) based on a rotatable central composite design (RCCD; half implementation), a pot experiment was performed in a natural-light greenhouse from June to Sept. 2014. The study assessed the interaction between irrigation volume (W) and nitrogen (N), phosphorus (P), and potassium (K) fertilizer rates on plant height, crown diameter, number of leaves, single leaf area, and fresh weight. Results showed that W had a significant positive effect on plant height, crown diameter, single leaf area, and fresh weight. Furthermore, P fertilization resulted in increased leaf number. Combined P and K fertilization reduced individual leaf area, whereas combined N and P fertilization reduced fresh weight. By analyzing the multiobjective decision-making model, we found that a combination of 100.2 mL water, 3.6 mmol·L−1 N, 0.10 mmol·L−1 P, and 1.2 mmol·L−1 K could be used to achieve optimum growth of gesnariad.
Cunquan Yuan, Zhiyi Qu, Huitang Pan, Tangren Cheng, Jia Wang and Qixiang Zhang
Heterostylous Primula forbesii is an important ornamental flower in China because of its long-lasting flowers and winter bloom. This study aimed to develop markers of expressed sequence tag–simple sequence repeats (EST-SSRs) that are associated with heterostyly and that can be used for molecular-assisted selective breeding in P. forbesii. We investigated 114,474 unigenes and identified 25,095 SSRs in P. forbesii. Dinucleotide repeats (46.14%), mononucleotide repeats (44.65%), and trinucleotide repeats (8.27%) were the most abundant SSRs. Among the 25,095 SSRs, 10,645 SSR primer pairs were successfully designed, of which 130 primer pairs were randomly selected for further amplification validation using eight accessions of P. forbesii; 98 pairs produced clear and stable polymerase chain reaction (PCR) products, and 28 pairs showed polymorphism. Bulked segregant analysis (BSA) was conducted for the F1 population with respect to thrum style and pin style by scanning 28 polymorphic SSR primer combinations. One SSR marker, c64326, linked to the heterostyly trait at a genetic distance of ≈3.70 cM was identified. The marker c64326 was further validated in two populations with an accuracy of 97.92% and 90.63%. The novel and linked EST-SSR markers can be valuable resources for genetic diversity analysis, mapping, and marker-assisted breeding in P. forbesii.
Jia-yi Wang, Jian-shuang Shen, Mengmeng Gu, Jia Wang, Tang-ren Cheng, Hui-tang Pan and Qi-xiang Zhang
Yellow-leafed cultivars usually do not grow as vigorous as their green-leafed counterparts, which affect their use in landscapes. To breed Forsythia cultivars with both yellow leaves and vigorous growth, crosses between F. ‘Courtaneur’ (♀) and Forsythia koreana ‘Suwon Gold’ (♂) were conducted, and 52 F1 hybrid progenies with different leaf colors (green, chartreuse, and yellow) were obtained. The progenies were categorized into three groups [Yellow Group (YG), Chartreuse Group (CG), and Green Group (GG)] based on leaf colors. The growth index (GI) and the number of branches and leaves of YG progenies were significantly lower at 2%, 35%, and 34% of GG progenies. As the leaves changed from green to chartreuse and to yellow, chlorophyll content, leaf thickness, and chlorophyll fluorescence parameters decreased and the chloroplast structures were disintegrated gradually, which influenced the leaf photosynthetic activity and led to weak growth. Compared with yellow-leafed progenies, the leaf chlorophyll content and leaf thickness of chartreuse-leafed progenies were significantly higher at 71% and 9%. The chloroplast structure of stroma lamella of chartreuse-leafed progenies was relatively intact. Carboxylation efficiency (CE), photochemical efficiency of PS II (F v/F m), and the number of branches and leaves of GG progenies were significantly higher than YG progenies; however, they have no significant difference with CG progenies. The results were promising for breeding new forsythia cultivars from moderate growth and chartreuse leaves.