Floral Stage Research Articles

CDNA Cloning and Expression Analysis of the Chalcone Synthases (CHS) in <i>Osmanthus fragrans</i>

In the flavonoid biosynthesis pathway, Chalcone synthase (CHS) is involved in the formation of the pigment and has been shown to be a rate-limiting enzyme for the synthesis of flavonoids. In this study, a PCR approach was used to clone a Chalcone synthases cDNA from flower of sweet osmanthus “Chenghong Dangui” and it was designated as OfCHS (O. fragrans, CHS). The cDNA was 1383 bp long and a coding sequence (CDS) of 1173 bp encoding a polypeptide of 391 amino acids with an estimated molecular mass of 39.9 kDa. The theoretical isoelectric point was 6.23. Phylogenetic analysis demonstrated that OfCHS clustered with Olea europaea, Solenostemon scutellarioides, Perilla frutescens, Antirrhinum majus and Digitalis lanata. We also detected the expression of OfCHS in different tissues in “Dangui” and in two cultivars with varied coloration, “Zi Yingui” and “Chenghong Dangui” at different floral stages using quantitative real-time PCR. We observed that OfCHS transcript was higher in leaves than in petals in “Dangui”. The transcripts of OfCHS in “Zi Yingui” petals were higher than those in “Dangui” at three stages especially at xianyan stage and there was no significant difference between the two cultivars in the full flowering stage. “Chenghong Dangui” has a relatively high anthocyanin content compared to “Zi Yingui”. The relative amount of anthocyanin of “Chenghong Dangui” initially increases, and then decreases during the bloom period. However, the expression of CHS is the highest at the initial flowering stage. These data suggest that the OfCHS does not play a key role in the accumulation of total flavonoid in this cultivar. These data could contribute to explain the different accumulation of flavonoids in petals of the two cultivars.

Low temperature-induced leaf senescence and the expression of senescence-related genes in the panicles of Litchi chinensis

Litchi is one of the most important subtropical evergreen fruit trees in Southern Asia. Litchi floral buds are a mix of axillary or apical panicle primordia, leaf primordia, and rudimentary leaves. Under usual winter and early spring conditions, the axillary panicle primordia prevail, and the rudimentary leaves abscise when low temperatures reach a certain threshold. The floral buds ultimately develop into pure panicles. Understanding the regulatory mechanism of rudimentary leaf senescence is of great importance for litchi flowering. In this study, litchi potted trees at the floral differentiation stage were treated with low and high temperatures in order to induce senescence or development of leaves. The microstructure of the petiole base of the rudimentary leaves was determined. The results show several layers of flattened cells forming in the abscission zone of the rudimentary leaves that were treated with low temperatures as well as an obvious boundary regarded as the abscission layer zone. We also determined the gene expression in the leaves with different developmental fate. The results show that the LcRboh, LcMC-1-like, and LcPirin genes were significantly induced in the rudimentary leaves treated with low temperatures, and the expression increased with the proceeding of senescence. The expression of the genes encoding class Ι β-1,3-glucanase and β-xylosidase also increased with the senescence, suggesting their possible involvement in the low temperature-induced senescence of the rudimentary leaves.

On the Presence of Uncommon Stylar Glandular Trichomes in Asteraceae: A Study in Kaunia R.M. King and H. Rob. (Oxylobinae, Eupatorieae)

Premise of research. Glands on the styles of Asteraceae are uncommon. They are reported in Kaunia for the first time. Glandular trichomes of plants are involved in the synthesis of secondary metabolites. In Kaunia, secondary compounds with medicinal cytotoxic, antitumor activity, and sensitizing capacity were previously identified. In this frame, a deeper knowledge of glandular trichomes could shed light on the productivity of medicinal substances in Kaunia. The potential taxonomic and ecological value of stylar glandular trichomes in Kaunia is also explored together with trichome secretion phases throughout the floral developmental stages.Methodology. Herbarium specimens and flowers fixed in FAA of 10 species of Kaunia were analyzed. Flowers in different developmental stages were studied to observe the secretory phases of stylar trichomes. A histochemical test was carried out to detect sesquiterpene lactones on stylar glandular trichomes.Pivotal results. Glandular trichomes were detected on the ventral face of style branches in eight species of Kaunia in all floral stages analyzed. The stylar trichomes observed were mostly found in secretory or early postsecretory phases in the most immature floral stages. The histochemical test in stylar trichomes was positive for sesquiterpene lactones.Conclusions. The presence of stylar glandular trichomes provides valuable new evidence for the identification and characterization of Kaunia. Within the Eupatorieae, the presence of stylar glandular trichomes could be interpreted as taxonomically valuable at the rank of genus. Our finding of sesquiterpene lactones is in agreement with the presence of Guaianolides and Germacranolides previously isolated in Kaunia. Apparently, the secretion of trichomes is completed before the opening of the corolla. The glands on styles of Kaunia could be the most internal barrier to herbivores that could flank outer floral structures during pollen and style development.

Responses of two strawberry cultivars to severe high temperature stress at different flower development stages

Prolonged heat stress negatively affects fruit set and fruit development in strawberry (Fragaria x ananassa Duch.), but the effect of a one-time severe heat stress at different stages of flower development is poorly understood. We hypothesized that strawberry is negatively affected by acute and severe heat stress but the response varies by floral development stage and by cultivar. Two Japanese cultivars, ‘Nyoho’ and ‘Toyonoka’, which were previously reported to differ in their tolerance to prolonged high temperatures, were heat stressed at 42°C for 4h at 12, 9, 6, 3, and 0days before anthesis (DBA) of the primary flower. Data on the percentage fruit set and fresh weight, diameter, and length of all developed fruits were then collected. In terms of fruit set from primary to tertiary flowers, two heat-sensitive floral development stages were observed in ‘Nyoho’: at 12 DBA and 0 DBA (or at anthesis). In ‘Toyonoka’, the heat-sensitive floral development stages were at 9 DBA and 0 DBA. Fresh weight and fruit size were larger in primary and secondary fruits of ‘Nyoho’ when the heat stress was applied during the earlier floral development stages but tertiary fruits were larger when heat stress was applied during the later floral development stages. For ‘Toyonoka’, the fresh weight and fruit size of primary fruits were smallest when the heat stress was applied at 9 and 0 DBA. These findings confirm our hypotheses and will be helpful in predicting yield losses in open-field strawberry production when a sudden heat wave occurs during the crop’s flowering and fruiting season from late winter to early summer.

Varietal variations in temperature response for hermaphrodite flower production and fruit set in mango (Mangifera indica L)

Mango produces hermaphrodite and staminate flowers on the same inflorescence and their ratio is influenced by genetic and environmental factors. Among the various environmental factors like temperature, rainfall, relative humidity, water stress and photoperiod, temperature is one of the most important factors influencing the production of hermaphrodite flowers. Therefore it is necessary to understand the influence of temperature in a locality on the flowering behaviour of different mango cultivars to select the most suitable type. To achieve this objective, the sex ratio of flowers (hermaphrodite/staminate) was recorded in panicles that emerged during the months of January, February and March and their relationship with mean minimum temperature during floral initiation stage was assessed for two years. Wide variability in flower sex ratio was observed among the cultivars and it ranged from 0.50 to 0.58 in cultivar Arka Aruna to 5.65 to 5.98 in cultivar Langra. The optimum, mean minimum temperature for maximum sex ratio in cultivars like Alphonso, Totapuri, Arka Aruna and Amrapali was 16.6 to 16.8°C and in cultivars Langra and Kensington it was 15.5 to 15.7°C. Fruit set was increased in the month of March followed by February in most of the cultivars indicating that the fruit set is influenced by both sex ratio and the higher temperature during flowering period.

The role of floral structure and biotic factors in determining the occurrence of florivorous thrips in a dystilous shrub

Elucidating the factors determining the occurrence of florivorous organisms is an essential step for comprehending arthropod–plant interactions, especially when considering florivores that use flowers/inflorescences as microhabitats. In this study, we characterize the interaction between florivorous thrips (Thysanoptera) and Palicourea rigida (Rubiaceae), a distylous hummingbird-pollinated shrub. We investigated the relative role of different factors in determining thrips occurrence in the flower and inflorescence microhabitats. Furthermore, we experimentally examined the protective role of corolla influencing thrips exploration of floral buds. Frankliniella musaeperda (Thripidae) was the only species recorded on P. rigida, feeding on floral tissue, pollen and nectar. Thrips occurrence was not related to distyly, but rather to floral stage. Open flowers presented the highest abundance of thrips, followed by senescent flowers and then buds. The experimental opening of buds translated in increased thrips occurrence, indicating that F. musaeperda manage to explore the microhabitat offered by the floral chamber, as long as there is an opening in the corolla. In inflorescences, thrips abundance was negatively related to the number of ants visiting extrafloral nectaries. We found that the marked difference between floral morphs of distylous plants is not necessarily reflected in the abundance of florivores. Thrips seek for floral cavities, preferentially those with fresh tissue, which may confer nutrient-rich food and protection. Buds also provide this; however, the enclosed petals are an effective barrier against F. musaeperda entrance. At inflorescence scale, presence of mutualistic ants in high numbers can drive away these flower-feeding insects. Despite the abundance of thrips in the flowers, there was no evidence of any functional relationship, either of pollination for flowers or of breeding for insects. We demonstrate here that in the flower/inflorescence microhabitat, structural and biotic factors play a key role in the exploitation and occupation by insect florivores.

Characterization of functional proteases from flowers of tea (Camellia sinensis) plants

Recently, functional compounds of flowers of tea (Camellia sinensis) have attracted increasing interest of researchers, whereas very less attention has been paid to functional enzymes in tea flowers. In the present study, tea flowers were found to contain proteases with strong proteolysis ability towards proteins from tea leaves. The crude proteases from tea flowers increased by 177% of total amino acids of tea infusion (protein:tea infusion, 1:3000), which was much higher than those of commercial proteases. LC–MS/MS analysis and protease inhibition experiments indicated that serine carboxypeptidase-like protein (SCLP) was one of major proteases in tea flowers. Recombinant CsSCLP was responsible for improvement of amino acids of tea infusion and CsSCLP was equivalently expressed in floral stages and most floral tissues. This study provides evidence of occurrences of proteases in tea flowers for the first time and an effective approach to improve contents of amino acids of tea infusion.

Morphogenetic and phenological changes in phenolic content of Hypericum leptophyllum, an endemic Turkish species

The present study was conducted to determine the morphogenetic and phenologic variations in the content of the main bioactive compounds in Hypericum leptophyllum Hochst., an endemic species from Turkish flora. Wild plants were harvested at five phenological stages: vegetative, floral budding, full flowering, fresh fruiting, and mature fruiting, and assayed for chemical content by HPLC. Results indicated that H. leptophyllum did not accumulate hypericin, pseudohypericin, hyperforin, and adhyperforin. The phenolic content in whole plants increased generally during plant phenology and higher accumulation levels were observed at flowering. Among the reproductive organs, amentoflavone, quercetin, avicularin, and hyperoside contents were the highest in floral buds (2.09, 2.18, 0.55, and 2.62 mg g−1 dry mass (DM), respectively). Leaves harvested at the floral budding stage produced the highest content of chlorogenic acid, neochlorogenic acid, caffeic acid, and isoquercetin (53.32, 6.25, 0.03, and 1.91 mg g−1 DM, respectively), while leaves of plants at the vegetative stage of plant development accumulated the highest content of 2,4-dihydroxybenzoic acid and quercitrin (0.29 and 0.54 mg g–1 DM, respectively). This is the first report of the presence of avicularin, rutin, (+)-catechin, (–)-epicatechin, amentoflavone, caffeic acid, neochlorogenic acid, and 2,4-dihydroxybenzoic acid in this species. Such data could be useful for the evaluation of new sources of bioactive compounds, as well as for increasing phytotaxonomic knowledge for the poorly studied Hypericum species.

Transcriptomic analysis of Arabidopsis overexpressing flowering locus T driven by a meristem-specific promoter that induces early flowering

Here we analyzed in leaves the effect of FT overexpression driven by meristem-specific KNAT1 gene homolog of Arabidopsis thaliana (Lincoln et al., 1994; Long et al., 1996) on the transcriptomic response during plant development. Our results demonstrated that meristematic FT overexpression generates a phenotype with an early flowering independent of photoperiod when compared with wild type (WT) plants. Arabidopsis FT-overexpressor lines (AtFTOE) did not show significant differences compared with WT lines neither in leaf number nor in rosette diameter up to day 21, when AtFTOE flowered. After this period AtFTOE plants started flower production and no new rosette leaves were produced. Additionally, WT plants continued on vegetative stage up to day 40, producing 12–14 rosette leaves before flowering. Transcriptomic analysis of rosette leaves studied by sequencing Illumina RNA-seq allowed us to determine the differential expression in mature leaf rosette of 3652 genes, being 626 of them up-regulated and 3026 down-regulated. Overexpressed genes related with flowering showed up-regulated transcription factors such as MADS-box that are known as flowering markers in meristem and which overexpression has been related with meristem identity preservation and the transition from vegetative to floral stage. Genes related with sugar transport have shown a higher demand of monosaccharides derived from the hydrolysis of sucrose to glucose and probably fructose, which can also be influenced by reproductive stage of AtFTOE plants.

Carbohydrate metabolism in floral structures of Lilium pumilum in different development stages

ABSTRACT: Lilium pumilum is a species that stands out in floriculture for presenting orange inflorescences that attract the consumer. This study thus aimed at characterizing the carbohydrate metabolism of floral structures of L. pumilum in different development stages. For this purpose, carbohydrate levels (total soluble sugars, reducing sugars, non-reducing sugars, and starch), at different floral stages (E0 - bud with no color; E1 - bud at early coloring; E2 - orange bud; E3 - open flower; E4 - senescent flower) were quantified after extraction with ethanol. Lilium pumilum flowers showed high energy potential during floral opening and senescence; total soluble sugars were the main carbohydrates present in the species, reducing with the floral development, and the same occurred with the non-reducing sugar and starch contents. The reducing-sugar content increased with the floral stages. Therefore, this species presents great mobilization of compounds, which are utilized in the production of energy that is employed in floral opening.

Pollinator responses to floral colour change, nectar, and scent promote reproductive fitness in Quisqualis indica (Combretaceae).

Floral colour change is visual signals for pollinators to avoid old flowers and increase pollination efficiency. Quisqualis indica flowers change colour from white to pink to red may be associated with a shift from moth to butterfly pollination. To test this hypothesis, we investigated Q. indica populations in Southwest China. Flowers secreted nectar continuously from the evening of anthesis until the following morning, then decreased gradually with floral colour change. The scent compounds in the three floral colour stages were similar; however, the scent composition was different, and the scent emission rate decreased from the white to red stage. Dichogamy in Q. indica prevents self-pollination and interference of male and female functions. Controlled pollinations demonstrated that this species is self-incompatible and needs pollinators for seed production. Different pollinators were attracted in each floral colour stage; mainly moths at night and bees and butterflies during the day. Observations of open-pollinated inflorescences showed that white flowers had a higher fruit set than pink or red flowers, indicating the high contribution of moths to reproductive success. We concluded that the nectar and scent secretion are related to floral colour change in Q. indica, in order to attract different pollinators and promote reproductive fitness.

APETALA2 and APETALA3 Genes Expression Profiling on Floral Development of Teak (Tectona grandis Linn f.)

eak (Tectona grandis Linn f.) is one of valuable timber species because of the beauty, strength and durability properties. Main problem in teak improvement program is reproductive biology, a low pollen viability, high embryo abortion, low pollination success and low fruit production. Investigation carried out at the teak improvement that flowering aspects is very important. Teak development program was facing difficulties due to the lack of information about the role of genes regulating flowering. APETALA2 (AP2) and APETALA3 (AP3) genes as member of the floral organ Identity. The role of teak flowering development predicted by up and down regulation of TgAP2 and TgAP3 genes. This study was conducted to determine the gene expression profile of TgAP2 and TgAP3 in the developmental stages of teak flowering. Materials and Methods: In this study, TgAP2 and TgAP3 genes in teak identified from NGS transcriptome data that is annotated with Solanum lycopersicum. The TgAP2 and TgAP3 genes activities on the several developmental stages of teak flowering tissues are done with QRT-PCR analysis using 18S RNA as gene reference. Results: The highest expression level of TgAP2 on the floral development stages of 4th lateral floral buds. The TgAP3 gene expression reached the highest level on the apical floral bud stage development. Vegetative shoot as control stages, TgAP2 gene expression level reached 9 times compared to the control and TgAP3 more than 12 times. The TgAP2 and TgAP3 genes expression profile has non-significant correlation on all floral development stages. Conclusion: The TgAP2 and TgAP3 genes expressed with different level in all stages of the teak flowering development. The TgAP2 and TgAP3 genes have non-significant correlation expression profile in role of teak floral development. These results indicate that the TgAP2 and TgAP3 expression profiles equivalent to the general pattern in the Arabidopsis model plant.

Cell division study in ‘pollen mother cells’ and ‘pollen grains’ of in vivo and ex vitro plants in Drimiopsis botryoides

The various normal and abnormal stages of meiosis and pollen mitosis of Drimiopsis botryoides are described, and a comparison between naturally propagated in vivo and tissue culture derived ex vitro plants in respect to their cytological behaviour presented. We also describe the floral morphology and investigate the relationship between the floral developmental stages and the progression of microgametogenesis. In total, 33 bivalents are observed in diakinesis, which indicate the diploid number 2n = 66 and this number is cross-checked by a haploid set of n = 33 chromosomes in pollen mitosis. Only 6.8% and 4.9% meiotic abnormalities were recorded on in vivo and ex vitro plants, respectively, which led to the formation of non-viable pollen. Finally, the microspores have to develop into tri-cellular male gametophyte. Only 0.2% pollen grains are found with a micro-nucleus. Though the higher pollen viability was recorded on both in vivo (89.3 ± 4.1%) and ex vitro (92.1 ± 4.6%) plants, but surprisingly the pollen germination rate is extremely low with 13.6 ± 1.74% and 21.3 ± 1.55%, respectively. The present study obviously enriches the cytological database of D. botryoides and may help future research on androgenesis and genetic improvement.

Susceptibility of Faba Bean (Vicia faba L.) to Heat Stress During Floral Development and Anthesis

Experiments were conducted over 2 years to quantify the response of faba bean (Vicia faba L.) to heat stress. Potted winter faba bean plants (cv. Wizard) were exposed to temperature treatments (18/10; 22/14; 26/18; 30/22; 34/26 °C day/night) for 5 days during floral development and anthesis. Developmental stages of all flowers were scored prior to stress, plants were grown in exclusion from insect pollinators to prevent pollen movement between flowers, and yield was harvested at an individual pod scale, enabling effects of heat stress to be investigated at a high resolution. Susceptibility to stress differed between floral stages; flowers were most affected during initial green‐bud stages. Yield and pollen germination of flowers present before stress showed threshold relationships to stress, with lethal temperatures (t 50) ˜28 °C and ~32 °C, while whole plant yield showed a linear negative relationship to stress with high plasticity in yield allocation, such that yield lost at lower nodes was partially compensated at higher nodal positions. Faba bean has many beneficial attributes for sustainable modern cropping systems but these results suggest that yield will be limited by projected climate change, necessitating the development of heat tolerant cultivars, or improved resilience by other mechanisms such as earlier flowering times.

Genomics and relative expression analysis identifies key genes associated with high female to male flower ratio in Jatropha curcas L.

Jatropha curcas, has been projected as a major source of biodiesel due to high seed oil content (42%). A major roadblock for commercialization of Jatropha-based biodiesel is low seed yield per inflorescence, which is affected by low female to male flower ratio (1:25-30). Molecular dissection of female flower development by analyzing genes involved in phase transitions and floral organ development is, therefore, crucial for increasing seed yield. Expression analysis of 42 genes implicated in floral organ development and sex determination was done at six floral developmental stages of a J. curcas genotype (IC561235) with inherently higher female to male flower ratio (1:8-10). Relative expression analysis of these genes was done on low ratio genotype. Genes TFL1, SUP, AP1, CRY2, CUC2, CKX1, TAA1 and PIN1 were associated with reproductive phase transition. Further, genes CUC2, TAA1, CKX1 and PIN1 were associated with female flowering while SUP and CRY2 in female flower transition. Relative expression of these genes with respect to low female flower ratio genotype showed up to ~7 folds increase in transcript abundance of SUP, TAA1, CRY2 and CKX1 genes in intermediate buds but not a significant increase (~1.25 folds) in female flowers, thereby suggesting that these genes possibly play a significant role in increased transition towards female flowering by promoting abortion of male flower primordia. The outcome of study has implications in feedstock improvement of J. curcas through functional validation and eventual utilization of key genes associated with female flowering.

The Application of BBCH Scale for Codification and Illustrations of the Floral Stages of Caribbean Fine Cacao Theobroma cacao L.

The Application of BBCH Scale for Codification and Illustrations of the Floral Stages of Caribbean Fine Cacao Theobroma cacao L.

Three FLOWERING LOCUS T-like genes function as potential florigens and mediate photoperiod response in sorghum.

Sorghum is a typical short-day (SD) plant and its use in grain or biomass production in temperate regions depends on its flowering time control, but the underlying molecular mechanism of floral transition in sorghum is poorly understood. Here we characterized sorghum FLOWERING LOCUS T (SbFT) genes to establish a molecular road map for mechanistic understanding. Out of 19 PEBP genes, SbFT1, SbFT8 and SbFT10 were identified as potential candidates for encoding florigens using multiple approaches. Phylogenetic analysis revealed that SbFT1 clusters with the rice Hd3a subclade, while SbFT8 and SbFT10 cluster with the maize ZCN8 subclade. These three genes are expressed in the leaf at the floral transition initiation stage, expressed early in grain sorghum genotypes but late in sweet and forage sorghum genotypes, induced by SD treatment in photoperiod-sensitive genotypes, cooperatively repressed by the classical sorghum maturity loci, interact with sorghum 14-3-3 proteins and activate flowering in transgenic Arabidopsis plants, suggesting florigenic potential in sorghum. SD induction of these three genes in sensitive genotypes is fully reversed by 1 wk of long-day treatment, and yet, some aspects of the SD treatment may still make a small contribution to flowering in long days, indicating a complex photoperiod response mediated by SbFT genes.

STIGMATIC RECEPTIVITY AND POLLEN VIABILITY OF Theobroma subincanum Mart.:FRUIT SPECIES FROM THE AMAZON REGION

ABSTRACT Theobroma subincanum (cupuí) is a fruit species native to the Amazon region. Fruits are enjoyed by local people and consumed both as fresh fruit such as juice, nectar or soft drinks. Reproductive biology studies provide contributions to conservation strategies and plant improvement. The present study aimed to analyze the stigmatic receptivity and pollen viability of T. subincanum. This study was developed in a forest fragment located in the urban perimeter of the municipality of Alta Floresta, MT. In the flowering period, flower buds or flowers were collected in seven different times, as follows: 10 pm, 02 am, 06 am, 10 am, 2 pm, 6 pm, and 10 pm. Pollen viability was estimated by reactive Alexander (1969) and stigmatic receptivity using 3% hydrogen peroxide. Pollen viability averages were submitted to analysis of variance, while stigmatic receptivity was analyzed by average percentage in each interval. Four floral stages were characterized based on flower opening, and from 6 am, fully opened flowers have already been found (stage IV). The percentage of pollen viability was not affected by collection times. In the stigmatic receptivity analysis, it was observed that in all floral stages, stigma was receptive; however, the highest percentages of stigmatic receptivity were found from 2 am to 10 am of the same day, which is the most propitious time for fertilization. Pollen collection of T. subincanum may be performed in any of schedules evaluated in this study, since it is held with high viability percentage.

Study on Petal-Sepal Mutant of Sunflower (<i>Helianthus annuus</i>) after Space Mutation

In this paper, we studied the morphological, histological and photosynthetic characteristics of the stably inherited sunflower petal-sepal mutant, and it was obtained by the space radiation-induced mutagenesis. Afterwards, we got following results: 1) The morphological characteristics represented that the inflorescence of petal-sepal mutant maintained the appearance and structure of capitulum, whereas no explicit tubular flower or ligulate flower was differentiated. 2) The histological characteristics revealed that the petal-sepal mutant only completed the inflorescence development and the differentiation of sepal primordia and inflorescence primordia, without entering the differentiation stage of tubular flower primordia, ligulate flower primordia, stamen primordia or pistil primordia. 3) The photosynthetic characteristics showed that the photosynthetic rate, transpiration and stomatal conductance of petal-sepal mutant were relatively weaker than the control plants. In the end, we concluded that the petal-sepal mutant of sunflower had only inflorescence differentiation, while several mutant genes were caused by radiation-induced mutation, which entered an infinitely recurrent development process rather than the floral differentiation stage. We also observed a few chloroplast structures in the paraffin section, combined with the results of photosynthetic characteristics of petal-sepal mutant, and then we believed that the inflorescence of petal-sepal mutant was involved in photosynthesis to accumulate energy for plant growth.

Validation of reference genes in pansy for accurate transcript normalization during its flowering stages

Quantitative real-time PCR (qRT-PCR) is a perfect method for rapid and accurate quantification of gene expression in different organs or at different development stages in plants. Suitable reference genes are important in this method. In order to obtain more accurate genes expression data in pansy (Viola × wittrockiana Gams.) flower, the expression stability of four housekeeping genes, -actin (ACT), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), tubulin (TUB) and 18S ribosomal proteins (18S) at seven different floral development stages in pansy were studied in this experiment. The results showed that TUB and 18S genes were the top two stable genes. Additionally, the expression pattern of VwbHLH1 gene was studied with the two most stable reference genesTUB and 18S, and the worst gene GAPDH respectively. The gene expression data were very different when various reference genes were applied. The present study proved that selection of reference genes was definitely important to obtain precise experimental data in qRT-PCR even for the same organ but at different development stages. This study would provide guidelines to obtain more accurate gene expression results for future molecular mechanism study in pansy.