Free-hand Sections Research Articles

Secretory structures and histochemistry of roots, stem, and leaves of medicinal plant Fibraurea tinctoria Lour.

Fibraurea tinctoria Lour. (Menispermaceae) is a medicinal plant used by many local tribes in Indonesia and other countries. This species has pharmacological potential, such as antimalaria, antibacterial, antitumor, antioxidant, anticancer, antidiabetic, anti-inflammatory, and hepato-protector. However, the secretory structures and sites of secondary metabolite accumulation still need to be studied. This study aimed to explore the secretory structures and reveal the accumulation sites of the main classes of secondary metabolites in the root, stem, and leaves of F. tinctoria. Histochemical analysis using various specific reagents was performed on freehand sections of the fresh organs to detect alkaloids, phenolics, terpenoids, and lipophilic compounds. Observations of secretory structures were carried out using light microscopy. The result showed that idioblast was the predominant secretory structure in the plant organs and contained all the chemical groups investigated, while laticifer accumulated alkaloids were found only in the stem. Some secondary metabolites were localized in the xylem, intercellular space, and cuticle layer. Crystals were observed in the root, stem, and leaf of this species. These results might be helpful as guide information for extracting the particular plant parts to obtain the desired secondary metabolites. It suggested that leaves are a potential alternative source of medicinal raw material of this species, in addition to the stem. RESEARCH HIGHLIGHTS: The first study on secretory structures of Fibraurea tinctoria. Secondary metabolites in F. tinctoria organs accumulate in specialized and unspecialized structures. The idioblast is the primary secretory structure in the organs of F. tinctoria spreading over the root, stem, and leaves.

NIGHTSHADE NECTAR: A MULTIFACETED DIVE INTO MEDICINAL, POLLEN, AND STEM ANATOMIES OF FIVE SOLANACEAE WONDERS

This comprehensive study delves into the intricate anatomy and reproductive biology of five prominent members of the Solanaceae family: Capsicum frutescens L., Solanum lycopersicum L., Solanum torvum Sw., Datura stramonium L., and Solanum melongena L. Through detailed anatomical examinations, including stem structure analyses, and investigations into pollen germination processes, this research elucidates the unique characteristics and adaptations of each species. Additionally, the study provides essential insights into the materials required for free-hand sectioning and pollen viability testing, facilitating further research in plant anatomy and reproductive physiology. With a focus on both morphological and physiological aspects, this work contributes to a deeper understanding of Solanaceae plant species and their ecological significance.

Echeveria Leaf Morpho-Anatomical Analysis and Its Implications for Environmental Stress Conditions

Echeveria, classified in the Crassulaceae family, possesses unique adaptive strategies with xeromorphic features to withstand semi-arid environments. The diversity and ecological adaptation of succulent plants offer valuable insights into addressing climate change challenges. In particular, the epidermis, hypodermis, vascular bundles arrangement, and stomata characteristics are commonly used to investigate light, humidity, temperature, and water availability adaptations. While leaf anatomical analysis is a common approach, limited studies have been conducted on Echeveria, especially among cultivars. To understand how succulents cope with environmental stress, leaf morpho-anatomical features were analyzed using the free-hand sectioning method with methanol fixation of fifteen Echeveria cultivars. The finding revealed a robust correlation between epidermis and hypodermis size (r = 0.362–0.729), and a positive association between leaf thickness and the epidermis (r = 0.362–0.536), suggesting implications for water storage. Most cultivars displayed a 3D vascular arrangement, with minor vascular bundles surrounding the main vascular bundle at the center, along with small stomata size, and low stomata frequency in the adaxial surface. Moreover, these cultivars grown under controlled conditions maintain their xeromorphic characteristics with the presence of epicuticular wax and thick and fully expanded small leaves. Likewise, the features of cultivars ultimately suggest that these succulents are tolerant to high temperatures and limited water supply. This study provides a fundamental understanding of Echeveria plants’ leaf anatomy and the correlation of their leaf structures toward environmental stress. Likewise, the methods and results of this study will serve as a benchmark for other research in related species.

Histochemical Investigation of Exotic Weed Lantana camara Linn

The histochemical Studies of leaves, Root and Stem of Lantena camara Linn.(Verbenaceae) in Maharashtra. It is a annual shrub grown widely in different places of India. The species has been investigated for the phytochemical, anti-oxidant and antimicrobial Potential. This paper encompasses a comprehensive review on histochemical and Biological aspects of Lantena camara L. For histochemical studies the freehand sections of leaves, stem and root were taken and treated with respective reagent in localize component, viz. Starch, protein, tannin, saponin, fat, and alkaloids in the tissues.
 Keywords: Histochemistry, Lantena, Alkaloids, Protein

STEM ANATOMY OF THREE SPECIES OF PINANGA FROM ASSAM, INDIA

The stem anatomy of the genus Pinanga Blume distributed to Assam of northeast India remains unattended. An anatomical study was conducted to address this research gap, where the plant samples to Pinanga were collected from different parts of Assam and identified as P. gracilis, P. griffithii, and P. sylvestris. The findings recorded anatomical variation among the species. Freehand sections of the stems were conducted, double-stained, quality specimens mounted, and suitable slides were observed under a light microscope. The types of epidermal cells and metaxylem vessels were variable among the studied species. In this study, vascular bundles with more than one metaxylem vessel were also recorded, which can be used for identification

Novel System for the Production of the Bioactive N-alkylamide ‘Spilanthol’ Through Somatic Embryogenesis in Acmella ciliata Kunth (Cass.)

Acmella ciliata Kunth (Cass.), a medicinally important plant in the family Asteraceae, has high commercial value because of its traditional phytomedicinal uses. The plant contains many phytochemicals like alkyl amides, alkaloids, tannins, saponins and flavonoids accountable for most of its pharmacological applications. The study presented here reports the callus culture and somatic embryogenesis of this plant thereby raising a novel system for the subsequent production of the N-alkyl amide ‘spilanthol’, the valuable secondary metabolite presents in it. Murashige and Skoog (MS) medium supplemented with auxins either alone or in combination with cytokinins were used for the induction and maturation of somatic embryos. MS medium supplemented with 2,4-D (0.5, 1.0 and 2.0 mg.L-1) produced black friable callus whereas, 1.0 mg.L-1 NAA in combination with 0.5 mg.L-1 BA induced white, slightly purple coloured friable callus which on further subculture to fresh medium induced somatic embryos that germinated into plantlets upon transfer to MS basal medium. The mode of regeneration via somatic embryogenesis was confirmed by histological analysis through free-hand sectioning and stereomicroscopic observation. The plantlets raised through somatic embryogenesis after a short hardening period, were found to acclimatise in the field at 83.33% efficiency and exhibited genetic uniformity with 96.6% similarity in the ISSR analysis. HPLC analysis of in vitro raised embryogenic callus showed 239.512 µg.g-1 spilanthol content which was comparatively higher than the mother plants (92.19 µg.g-1). The bioproduction of the N-alkylamide ‘spilanthol’ through embryogenic callus can be extended for the scale-up production of this bioactive compound using bioreactor technology for the formulation of phytodrugs.

Anatomical Study for Vegetative Organs of Pelargonium graveolens L.'Hér Cultivated in Iraq

The current investigation concluded an anatomical study of the cultivated species Pelargonium graveolens L. 'Hér belong to the Geraniaceae family and which cultivated as an ornamental plant, fresh specimen were collected from different region in Baghdad ,the study vegetative parts (leaf, stem and petiole) in details ,the upper and lower epidermis of leaf and epidermis of stem also studied, ordinary epidermal cells and stomatal complexes studied in term of shapes and dimensions ,the result showed that stomatal in leaf and stem is anomocytic type, the cross sections of the vegetative parts prepared by microtome and free hand sectioning. The tissues of these parts include the number and the type of layers and thickness of each one , the result showed the presence of druses crystals in parenchyma cells of the petiole, stem and leaf. Vistur type of this organs also studied and it comprises of glandular and non-glandular trichomes which occur in different types and lengths.

Identification and characterization of the gene BraANS.A03 associated with purple leaf color in pak choi (Brassica rapa L. ssp. chinensis).

BraANS.A3 was the key gene controlling purple leaf color in pak choi, and two short fragments of promoter region in green pak choi might be interfering its normal expression. Pak choi (B. rapa L. ssp. chinensis) is an influential and important vegetable with green, yellow, or purple leaves that is cultivated worldwide. The purple leaves are rich in anthocyanins, but the underlying genetics and evolution have yet to be extensively studied. Free-hand sections of the purple leaves indicated that anthocyanins mainly accumulate throughout the adaxial and abaxial epidermal leaf cells. Segregation analyses of an F2 population of a B. rapa ssp. chinensis L. purple leaf mutant ZBC indicated that the purple trait is controlled by an incompletely dominant nuclear gene. Bulked segregant analysis (BSA) showed that the key genes controlling the trait were between 24.25 and 38.10Mb on chromosome A03 of B. rapa. From the annotated genes, only BraA03g050560.3C, homologous to Arabidopsis AtANS, was related to the anthocyanin synthesis pathway. Genome annotation results and transcriptional sequencing analyses revealed that the BraANS.A3 gene was involved in the purple leaf trait. qRT-PCR analyses showed that BraANS.A3 was highly upregulated in ZBC but hardly expressed in the leaves of an inbred homozygous line of B. campestris ssp. chinensis L. green leaf mutant WTC, indicating that BraANS.A3 played a key role catalyzing anthocyanin synthesis in ZBC. Full-length sequence alignment of BraANS.A3 in WTC and ZBC showed that it was highly conserved in the gene region, with significant variation in the promoter region. In particular, the insertion of two short fragments of the promoter region in WTC may interfere with its normal expression. The promoter regions of ANS in six Brassica species all had multiple cis-elements involved in responses to abscisic acid, light, and stress, suggesting that ANS may be involved in multiple metabolic pathways or biological processes. Protein-protein interactions predicted that BraANS.A3 interacts with virtually all catalytic proteins in the anthocyanin synthesis pathway and has a strong relationship with Transparent Testa 8 (TT8). These results suggest that BraANS.A3 promotes anthocyanin accumulation in purple pak choi and provide new insights into the functional analysis of anthocyanin-related genes in Chinese cabbage and transcriptional regulatory networks.

Physiological and anatomical response of rice (Oryza sativa L.) ‘Hom Mali Daeng’ at different salinity stress levels

Abstract Soil salinity is a severe global stressor causing adverse impacts on irrigated land and drastically reducing crop yields, especially in rice, an important economic crop of Thailand. In this study, the impacts of salt stress on the anatomical and physiological features of 28-day-old rice (Oryza sativa L.) ‘Hom Mali Daeng’ were determined. Various NaCl concentrations (0, 50, 100, 150, and 200 mM) were applied every 2 days, with watering for 2 weeks. The results revealed that salinity stress inhibited the growth of rice. Leaf number, root size, fresh weight, and dry weight were significantly reduced. The electrolyte leakage percentage and malondialdehyde (MDA) content increased after treatment with high NaCl concentrations, while the SPAD unit and chlorophyll content were not significantly different between the control and NaCl treatments. Leaf anatomy changes were studied using freehand section and peeling techniques after salinity stress treatment. Lamina thickness in all treatments decreased, while cell wall and cuticle thickness increased. Stomatal density in all treatments significantly increased. Major vascular bundle, vessel, and phloem area of the 100 mM NaCl treatment were different when compared with the control and other treatments. The results provide information about the physiological and anatomical adaptation of ‘Hom Mali Daeng’ rice, which will be useful for further research in this and other rice cultivars.

Genetic and transcriptome analysis of a cotton leaf variegation mutant

In eukaryotic photosynthetic organisms, chloroplast is not only a site for photosynthesis, but it also have a vital role in signal transduction mechanisms. Plants exhibit various colors in nature with various mutants induced by EMS, whose traits are regulated by developmental and environmental factors, making them ideal for studying the regulation of chloroplast development. In this study, the cotton leaf variegated mutant (VAR) induced by EMS was used for this experiment. Genetic analysis revealed that VAR phenotype was a dominant mutation and by performing freehand section inspection, it was noticed that the vascular bundles of VAR were smaller. Chloroplast ultrastructure showed that the stacking of grana thylakoid was thinner and the starch granules were increased significantly in VAR comparedto wild type (WT). Transcriptome analysis found that the KEGG was enriched in photosynthesis pathway, and GO was abundant in zinc ion transmembrane transport, electron transporter and cation binding terms. In addition, GhFTSH5 expression in VAR was significantly higher than WT and the promoter sequence of GhFTSH5 had differences. The results showed that the VAR plant had altered GhFTSH5 expression and disrupted chloroplast structure, which in turn affects plant photosynthesis. More importantly, this study lays a foundation for further analyzing molecular mechanism of cotton variegated phenotypes.

Structural modifications in Bermuda grass [Cynodon dactylon (L.) Pers.] ecotypes for adaptation to environmental heterogeneity.

It is well known that different ecotypes adopt different mechanisms to survive under environmental stress conditions. In this regard, each ecotype showed different type of modifications for their existence in a specific habitat that reflects to their ecological success. Here, differently adapted ecotypes of Bermuda grass [Cynodon dactylon (L.) Pers.] were collected to evaluate their differential structural and functional modifications that are specific to cope with environmental stress conditions. The soil that adheres ecotypes roots were highly saline in case of DF-SD (Derawar Fort-Saline Desert), UL-HS (Ucchali Lake-Hyper Saline) and G-SSA (Gatwala-Saline Semiarid) ecotypes. Soils of S- HS (Sahianwala-Hyper Saline), S-SW (Sahianwala-Saline Wetland) and PA-RF (Pakka Anna-Reclaimed Field) were basic (pH 9 to 10). Soils of UL-HS and PA- HS (Pakka Anna-Hyper Saline), KKL-S (Kalar Kahar Lake-Saline), BG-NS (Botanic Garden-Non Saline) and G-SSA were rich in organic matter, and soil of BG-NS and DF-SD were rich in minerals. Anatomical modifications were performed by using the free hand sectioning technique and light microscopy. DF-SD is one of the best ecotypes which showed anatomical modifications to cope with environmental changes. These modifications included stem cross-sectional area and leaf sheath thickness that contribute towards water storage, vascular tissues for proficient translocation of solutes and trichomes that provide resistance to water loss. On the other hand, sclerification in root is the only notable modification in the Gatwala Saline Semiarid (G-SSA) ecotype from saline arid habitat where rainfall is not as low as in the Cholistan Desert. Two ecotypes from hyper-saline wetlands, UL-HS and KL-HS showed increased number and size of vascular tissue, central cavity and sclerification in stem which are important for solutes conduction, water loss and salts bulk movement, respectively. The ecotype from reclaimed site was not much different from its counterpart from hyper-saline dryland. Overall, anatomical modifications to maintain water conservation are key mechanisms that have been identified as mediating stress tolerance in C. dactylon ecotypes.

Comparative macro-microscopic studies of three wild Solanum species (S. melongena L., S. incanum L. and S. virginianum L.)

Background: The traditional medicinal system comprises the usage of innumerable plant species for the treatment of various ailments. Solanum, a potent medicinal genus, finds vast description in the various traditional medicinal systems world over. The effectiveness of a herbal drug relies on the usage of correct botanical sources. In this scenario of growing importance of traditional medicines and excessive loss of habitats of various medicinal plants, the procurement of genuine herbals is a challenging issue. Many related species of Solanum distributed over the world find their usage in many medicinal preparations, but the authenticity of the herbals is not promised. The identification and standardization of crude herbal drugs can be assured by the definite morphological and anatomical characterization. Materials and Methods: Macroscopical characters of all three samples collected from Attur Chat village in Salem district were observed and recorded using Nikon Digital camera D5600 and ZEISS Discovery V8 stereo microscope. The samples were preserved in formalin acetic acid, free hand sectioning was done to reveal the anatomy, and photographs were taken using Axiolab 5 trinocular microscope fitted with Axiocam 208 camera. Results: Macro-microscopic features of three important Solanum species viz S. melongena L., S. incanum L., and S. virginianum L. were characterized by morphology and anatomy studies. The stem was terete (S. incanum), flattened (S. melongena), and cylindrical (S. virginianum) in the species. The leaves were large and elliptic in S. melongena, simple ovate in S. incanum, and ovate and brittle in S. virginianum. Anatomically in the stem of S. virginianum, glandular trichomes were observed, which were absent in the other two species. In the case of leaves, the trichome type and density were varying in the three species, and trichomes were present only in the midrib of S. melongena. Variations observed in petiole and fruits were also recorded. Conclusion: The macro-microscopic characterization carried out will be worthwhile in judging the authenticity of the plants and differentiating these species from one another.

Identification of genetic loci for flag-leaf-related traits in wheat (Triticum aestivum L.) and their effects on grain yield.

Flag-leaf-related traits including length (FLL), width (FLW), area (FLA), thickness (FLT), and volume (FLV) of flag leaves are the most important determinants of plant architecture and yield in wheat. Understanding the genetic basis of these traits could accelerate the breeding of high yield wheat varieties. In this study, we constructed a doubled haploid (DH) population and analyzed flag-leaf-related traits in five experimental locations/years using the wheat 90K single-nucleotide polymorphism array. It’s worth noting that a novel method was used to measure FLT and FLV easily. Leaf thickness at two-thirds of the leaf length from tip to collar represented the average leaf thickness as measured with freehand sections and was used to calculate the leaf volume. In addition, flag-leaf-related traits showed positive correlations with yield related traits under two different water regimes. A total of 79 quantitative trait loci (QTL) controlling the five traits were detected among all chromosomes except 4D and 5A, explaining 3.09–14.52% of the phenotypic variation. Among them, 15 stable QTL were identified in more than three environments, including two major QTL for FLT, six for FLW, three for FLA, two for FLT and two for FLV. DH lines with positive alleles at both QTL regions had an average FLL (9.90%), FLW (32.87%), FLT (6.62%), FLA (18.47%), and FLV (20.87%) greater than lines with contrasting alleles. QFLT-2B, QFLV-2A, and QFLV-7D were co-located with yield-related traits. The 15 QTL were validated by tightly linked kompetitive allele specific PCR (KASP) markers in a recombinant inbred line (RIL) population derived from a different cross. QFLL-4A, QFLW-4B, QFLA-5D.1, QFLA-7A, QFLA-7D.1, QFLT-2B, QFLT-6A, QFLV-2A, and QFLV-7D are likely novel loci. These results provide a better understanding of the genetic basis underlying flag-leaf-related traits. Also, target regions for fine mapping and marker-assisted selection were identified and these will be valuable for breeding high yielding bread wheat.

Morphological and molecular characterization of some popular cultivated mushrooms of India

Morphological and molecular characterization of some popular cultivated mushrooms of IndiaSanjeev Kumar1*, Anand Sagar2, Shwet Kamal3 and Jagriti Rana41. Associate professor, Govt. College Bilaspur.2. Dean of Life Sciences, H.P. Univ.. Shimla. 3. Principal Scientist, DMR Solan.4. PhD research Scholar, Department of Biosciences, H.P. Univ.Shimla.* Corresponding author, E-mail: sanjeev5112@gmail.com +919418045021 AbstractThe mushroom cultivation is a highly lucrative agribusiness to produce nutritious food and earn money in short space and time. The present studies were conducted to analyze morphological and molecular characteristics of five cultivated strains of mushrooms Agaricus bisporus (NBS-5), Calocybe indica (DMRO-302), Flammulina velutipes (DMRO-253), Pleurotus florida (DMRP-136) and Volvariella volvacea (DMRO-484). The fruiting bodies were procured from ICAR-Directorate of Mushroom Research, Solan, HP (India) and screened for morphological and molecular details. Macroscopic features like shape, size and colour of stipe and pileus were recorded. Microscopic details were worked out by cutting free hand sections of material and observed under research microscope and scanning electron microscope. Molecular identification was done through internal transcribed spacer (ITS) regions of ribosomal DNA sequencing. The nucleotide sequences of each sample were analyzed for Basic local Alignment Search Tool (BLAST) search program in US National Centre for Biotechnology Information (NCBI) for identification. The DNA sequences obtained in this study were submitted in the Gen Bank database at NCBI with accession numbers MW718994.1 (A. bisporus), MW714627.1 (C. indica), MW719474.1 (F. velutipes), MW719254.1 (P. florida) and MW720603.1 (V. volvacea).Keywords: Mushroom, Molecular identification, ITS, rRNA, Nucleotide sequencing, Gen Bank database

Morphological, Anatomical, and Palynological Studies of Lasianthera africana P. Beauv (Stemonuraceae) from some Parts of Southern Nigeria

This study examined Lasianthera africana P. Beauv based on, anatomical and palynological parameters. The anatomical studies were carried out with freehand sectioning. The species is hypostomatic, the epidermal cells are are irregular, with undulating anticlinal walls. The stomata types are anomocytic, and anisocytic. The midrib has bicollateral vascular bundles and the petiolar vascular bundle is arced with rib traces on both sides of the wings. Calcium oxalate and tannin occurred in the lamina, midrib, petiole and stem. The pollen grains are smooth, intectate, with thick exine, with tricolporate aperture, and with an equatorial diameter of 21.5-25.8 µm. These features are diagnostic and confirm the placement of this species in Stemonuraceae.

Histochemical Investigation and Localization of Secondary Metabolites of Acalypha paniculata Miq.

The present study was focused to localize the bioactive molecules histochemically in leaves, stem and root of Acalypha paniculata Miq. Free hand sections of leaf, stem and root was obtained and treated with respective specific reagents to localize constituents such as starch, protein, carbohydrate, total lipids, phenols, tannins, flavonoids, saponins and alkaloids. The medicinal properties of the study plant Acalypha paniculata is due to the presence of secondary metabolites, like phenols, tannins, flavonoids, saponins and alkaloids.

Leaf Anatomical Adaptation Under Early Drought Stress of Sugarcane Cultivars – KKU-1999-02 and KKU-1999-03

Abstract Anatomical adaptation is an important feature that allows plants to mitigate drought stress. A comparative leaf anatomy of two drought-tolerant sugarcane cultivars, KKU-1999-02 and KKU-1999-03, was studied in early drought stress between 30 and 90 days after planting using peeling and freehand sectioning methods. KKU-1999-02 and KKU-1999-03 showed different anatomical adaptation features, such as increase in cuticle thickness, bulliform cell size, vascular bundle, and stomatal density, and decreases in leaf thickness and stomatal size. KKU-1999-02 showed more remarkable anatomical changes than KKU-1999-03. The results provide important information that can be applied in combination with other agronomic traits in sugarcane breeding programs to expand the adaptation devices of tolerant cultivars under preliminary drought stress.

Histochemical Test of root, petiole and leaf of Kelembak (Rheum officinale Baill.)

Kelembak (Rheum officinale Baill.) is perennial shrub with pharmacological activities, and well-known as one of traditional herbal medicine used widely in the world. The phytochemical study, as well as histochemical test, is pivotal to determine the potency of herbs used for disease treatment. Based on its secondary metabolite content. Histochemical test of R.officinale vegetative organs had not been reported. This study aims to determine the content of compounds in the vegetative organs of kelembak and provide information on the anatomical position of each compound in root, petiole and leaf. The preparation of histochemical test used freehand section method. Each sample was made in three replications for this test. A total of four secondary metabolites were examined in this study, i.e. alkaloid, tannin, lipid and flavonoid. The positive content of each compound was indicated by the coloration change. The specimens were then observed and photographed using light microscope. The results showed that the root, petiole and leaf showed positive content of alkaloids, tannins, lipids and flavonoids. However, the anatomical position of each secondary metabolites are different among root, petiole and leaf. The results of this study provide the first information of histochemistry vegetative organs of Rheum officinale.

Contribution to the identification of the species Myrcia hatschbachii D. Legrand (Myrtaceae): anatomical and histochemical analyses

The genus Myrcia is used in folk medicine to treat diabetes. The plants used in folk medicine require morphological and anatomical references to attest to its authenticity. This is the first report of the microscopic study of Myrcia hatschbachii. In this sense, the aim of the present study was to describe the anatomical characteristics, in order to contribute to the species' identification. For anatomical and surface analyses free-hand sections were prepared and observed using optical microscopy, simultaneously some materials were processed and observed under scanning electron microscopy. In addition, histochemical tests were performed. The anatomical features described here correspond with previously reported features found in other species of the genus Myrcia. Furthermore, the following anatomical markers were observed in Myrcia hatschbachii: druses in the spongy parenchyma, concave-convex midrib, heart-shaped petiole, oval-shaped stem, C-shaped vascular bundle in the midrib and open arc shape with invaginated ends in the petiole; phenolic compounds in the phloem and lignified cells in the pith. The results of anatomical and histochemical analyses provide micromorphological and microchemical features that can help in the taxonomy and proper identification of the species.

Frequent gain- and loss-of-function mutations of the BjMYB113 gene accounted for leaf color variation in Brassica juncea

BackgroundMustard (Brassica juncea) is an important economic vegetable, and some cultivars have purple leaves and accumulate more anthocyanins than the green. The genetic and evolution of purple trait in mustard has not been well studied.ResultIn this study, free-hand sections and metabolomics showed that the purple leaves of mustard accumulated more anthocyanins than green ones. The gene controlling purple leaves in mustard, Mustard Purple Leaves (MPL), was genetically mapped and a MYB113-like homolog was identified as the candidate gene. We identified three alleles of the MYB113-like gene, BjMYB113a from a purple cultivar, BjMYB113b and BjMYB113c from green cultivars. A total of 45 single nucleotide polymorphisms (SNPs) and 8 InDels were found between the promoter sequences of the purple allele BjMYB113a and the green allele BjMYB113b. On the other hand, the only sequence variation between the purple allele BjMYB113a and the green allele BjMYB113c is an insertion of 1,033-bp fragment in the 3’region of BjMYB113c. Transgenic assay and promoter activity studies showed that the polymorphism in the promoter region was responsible for the up-regulation of the purple allele BjMYB113a and high accumulation of anthocyanin in the purple cultivar. The up-regulation of BjMYB113a increased the expression of genes in the anthocyanin biosynthesis pathway including BjCHS, BjF3H, BjF3’H, BjDFR, BjANS and BjUGFT, and consequently led to high accumulation of anthocyanin. However, the up-regulation of BjMYB113 was compromised by the insertion of 1,033-bp in 3’region of the allele BjMYB113c.ConclusionsOur results contribute to a better understanding of the genetics and evolution of the BjMYB113 gene controlling purple leaves and provide useful information for further breeding programs of mustard.