Discovery and validation of candidate genes for grain iron and zinc metabolism in pearl millet [Pennisetum glaucum (L.) R. Br.].

Pearl millet is an important crop for alleviating micronutrient malnutrition through genomics-assisted breeding for grain Fe (GFeC) and Zn (GZnC) content. In this study, we identified candidate genes related to iron (Fe) and zinc (Zn) metabolism through gene expression analysis and correlated it with known QTL regions for GFeC/GZnC. From a total of 114 Fe and Zn metabolism-related genes that were selected from the related crop species, we studied 29 genes. Different developmental stages exhibited tissue and stage-specific expressions for Fe and Zn metabolism genes in parents contrasting for GFeC and GZnC. Results revealed that PglZIP, PglNRAMP and PglFER gene families were candidates for GFeC and GZnC. Ferritin-like gene, PglFER1 may be the potential candidate gene for GFeC. Promoter analysis revealed Fe and Zn deficiency, hormone, metal-responsive, and salt-regulated elements. Genomic regions underlying GFeC and GZnC were validated by annotating major QTL regions for grain Fe and Zn. Interestingly, PglZIP and PglNRAMP gene families were found common with a previously reported linkage group 7 major QTL region for GFeC and GZnC. The study provides insights into the foundation for functional dissection of different Fe and Zn metabolism genes homologs and their subsequent use in pearl millet molecular breeding programs globally.

Transcriptome Analysis to Shed Light on the Molecular Mechanisms of Early Responses to Cadmium in Roots and Leaves of King Grass (Pennisetum americanum × P. purpureum).

King grass, a hybrid grass between pearl millet and elephant grass, has many excellent characteristics such as high biomass yield, great stress tolerance, and enormous economic and ecological value, which makes it ideal for development of phytoremediation. At present, the physiological and molecular response of king grass to cadmium (Cd) stress is poorly understood. Transcriptome analysis of early response (3 h and 24 h) of king grass leaves and roots to high level Cd (100 µM) has been investigated and has shed light on the molecular mechanism underlying Cd stress response in this hybrid grass. Our comparative transcriptome analysis demonstrated that in combat with Cd stress, king grass roots have activated the glutathione metabolism pathway by up-regulating glutathione S-transferases (GSTs) which are a multifunctional family of phase II enzymes that detoxify a variety of environmental chemicals, reactive intermediates, and secondary products of oxidative damages. In roots, early inductions of phenylpropanoid biosynthesis and phenylalanine metabolism pathways were observed to be enriched in differentially expressed genes (DEGs). Meanwhile, oxidoreductase activities were significantly enriched in the first 3 h to bestow the plant cells with resistance to oxidative stress. We also found that transporter activities and jasmonic acid (JA)-signaling might be activated by Cd in king grass. Our study provided the first-hand information on genome-wide transcriptome profiling of king grass and novel insights on phytoremediation.

Effect of EDTA and NTA on cadmium distribution and translocation in Pennisetum purpureum Schum cv. Mott.

The primary objective of this research was to investigate the cadmium (Cd) distribution in Pennisetum purpurem (Napier grass) in the presence of 30 mg/L of Cd and different types and concentrations of chelating agents (ethylenediaminetetraacetic acid disodium dihydrate (EDTA), nitrilotriacetic acid (NTA), and EDTA-NTA mixtures). Plant samples were collected every 15 d during a 105-d experimental period. Accumulation of Cd in each part of the plant was determined using atomic absorption spectrometer (AAS), and the distribution of Cd was determined by laser ablation inductively coupled plasma mass spectrometer (LA-ICP-MS) and synchrotron radiation micro X-ray fluorescence (SR-micro-XRF). The highest concentrations of Cd accumulation of 889 ± 53 mg kg-1 in the underground part (roots) and 265 ± 26 mg kg-1 in the aboveground part (stems and leaves) in the presence of 1:1 M ratio of Cd:EDTA after 30 d of exposure were observed. Plants grown in the presence of either NTA or EDTA-NTA mixtures showed significant lower Cd accumulation levels. The LA-ICP-MS analysis showed that Cd was primarily accumulated in the aboveground part (stems and leaves), especially in the xylem and intercalary meristem. In addition, translocation factor was very low. Thus, P. purpurem could be considered as a candidate plant for cadmium phytostabilization.

Myosin XI is associated with fitness and adaptation to aridity in wild pearl millet.

Phenotypic changes in plants can be observed along many environmental gradients and are determined by both environmental and genetic factors. The identification of alleles associated with phenotypic variations is a rapidly developing area of research. We studied the genetic basis of phenotypic variations in 11 populations of wild pearl millet (Pennisetum glaucum) on two North-South aridity gradients, one in Niger and one in Mali. Most of the 11 phenotypic traits assessed in a common garden experiment varied between the populations studied. Moreover, the size of the inflorescence, the number of flowers and aboveground dry mass co-varied positively with a decrease in rainfall. To decipher the genetic basis of these phenotypes, we used an association mapping strategy with a mixed model. We found two SNPs on the same myosin XI contig significantly associated with variations in the average number of flowers. Both the allele frequency of the two SNPs and the average number of flowers co-varied with the rainfall gradient on the two gradients. Interestingly, this gene was also a target of selection during domestication. The Myosin XI gene is thus a good candidate for fitness-related adaptation in wild populations.

Silagem de capim-elefante adicionada de casca de maracujá / Elephant grass silage added to passion fruit's peel

Avaliaram-se-se as características fermentativas e químico-bromatológicas da silagem de capim-elefante contendo diferentes níveis de inclusão do coproduto da extração da polpa do maracujá, a casca de maracujá (CM). Utilizaram-se níveis crescentes de adição desse coproduto (0,0%; 12,5%; 25,0%; 37,5% e 50,0%) em relação à matéria natural do capim-elefante durante a ensilagem. As silagens foram obtidas a partir do corte da forrageira aos 150 dias de idade. O material foi ensilado em silos laboratoriais e, após 180 dias, os silos foram abertos e coletaram-se amostras para a determinação dos teores de matéria seca (MS); carboidratos solúveis (CHOS); potencial hidrogeniônico (pH); nitrogênio amoniacal em porcentagem do nitrogênio total (NNH3/NT); extrato etéreo (EE); proteína bruta (PB); proteína insolúvel em detergente neutro em porcentagem do nitrogênio total (PIDN/NT); proteína insolúvel em detergente ácido em porcentagem do nitrogênio total (PIDA/NT); fibra em detergente neutro (FDN); fibra em detergente ácido (FDA); celulose (CEL); lignina (LIG); nitrogênio insolúvel em detergente neutro (NIDN); nitrogênio insolúvel em detergente ácido (NIDA) e digestibilidade in vitro da matéria seca (DIVMS). Para a avaliação do efeito dos tratamentos, utilizou-se delineamento experimental inteiramente ao acaso, com quatro repetições, executando-se o estudo de regressão para cada variável analisada (P<0,05). De acordo com as equações de regressão, para as características fermentativas, as silagens podem ser consideradas de muito boa qualidade. Quanto às frações fibrosas da silagem, a inclusão de níveis crescentes desse coproduto contribuiu para diminuí-las, o que resultou em maior DIVMS. Os parâmetros analisados permitem concluir que a inclusão da CM in natura durante a ensilagem do capim-elefante é recomendada em todos os níveis avaliados.

We evaluated the fermentative, chemical and qualitative characteristics of elephant grass silage containing different levels of inclusion of coproduct extraction of the passion fruit pulp in nature, passion fruit peel (CM). We used increasing levels of addition of this coproduct (0.0, 12.5, 25.0, 37.5 and 50.0%) in relation to natural field of elephant grass during ensiling. The silages were obtained from the cut forage at 150 days of age. The material was ensiled in laboratory silos, and after 180 days, the silos were opened and samples were collected for determination of dry matter (DM); soluble carbohydrates (CHOS); hydrogen potential (pH); ammonia nitrogen in percentage of total nitrogen (NNH3/NT); ether extract (EE); crude protein (CP); neutral detergent insoluble protein as a percentage of total nitrogen (NDIP/NT); acid detergent insoluble protein as a percentage of total nitrogen (PIDA/NT); neutral detergent fiber (NDF); acid detergent fiber (ADF); cellulose (CEL); lignin (LIG); neutral detergent insoluble nitrogen (NDIN); acid detergent insoluble nitrogen (ADIN) and in vitro digestibility of dry matter (IVDDM). To assess the effect of the treatments we used a completely randomized design with four replications, running the regression analysis for each variable analyzed (P<0.05). According to the regression equations for the fermentative characteristics, silage can be considered very good quality. As for fiber digestibility of silage, the increasing levels of this coproduct contributed to decrease it, which resulted in higher IVDDM. The parameters analyzed showed that the inclusion of CM in nature with elephant grass silage is recommended at all levels evaluated.

Cloning and molecular characterization of a gene encoding late embryogenesis abundant protein from Pennisetum glaucum: protection against abiotic stresses.

Late embryogenesis abundant (LEA) protein family is a large protein family that protects other proteins from aggregation due to desiccation or osmotic stresses. A cDNA clone encoding a group 7 late embryogenesis abundant protein, termed PgLEA, was isolated from Pennisetum glaucum by screening a heat stress cDNA library. PgLEA cDNA encodes a 176 amino acid polypeptide with a predicted molecular mass of 19.21 kDa and an estimated isoelectric point of 7.77. PgLEA shares 70-74% sequence identity with other plant homologs. Phylogenetic analysis revealed that PgLEA is evolutionarily close to the LEA 7 group. Recombinant PgLEA protein expressed in Escherichia coli possessed in vitro chaperone activity and protected PgLEA-producing bacteria from damage caused by heat and salinity. Positive correlation existed between differentially up-regulated PgLEA transcript levels and the duration and intensity of different environmental stresses. In silico analysis of the promoter sequence of PgLEA revealed the presence of a distinct set of cis-elements and transcription factor binding sites. Transcript induction data, the presence of several putative stress-responsive transcription factor binding sites in the promoter region of PgLEA, the in vitro chaperone activity of this protein and its protective effect against heat and salt damage in E. coli suggest a role in conferring abiotic stress tolerance in plants.

Efeito de diferentes herbicidas, doses e volume de calda na dessecação de milheto [Pennisetum Glaucum (L. Leek)] / Effect of different herbicides, doses and spray volume in the chemical management of pearl millet [Pennisetum Glaucum (L. Leek)]

Objetivou-se nesse trabalho avaliar a eficácia de diferentes herbicidas aplicados em pósemergência na de secação do milheto com diferentes doses e volumes de calda de aplicação. Para a dessecação do milheto foram utilizadas doses de 0, 25, 50 e 100% dos herbicidas glyphosate (1.440 g ha-1), em três formulações comerciais (Round up Original, Round up Transorb e Round up WG), paraquat + diuron (500 + 250 g ha-1) e amônio-glufosinato (800 g ha-1). Em todos os tratamentos foram utilizados dois volumes de aplicação (200 e 400 L ha-1). Manteve-se uma testemunha sem aplicação de herbicidas. Os efeitos dos tratamentos foram avaliados visualmente aos 3, 7, 14, 21 e 28 dias após aplicação. Ao final das avaliações foi determinada a massa seca das plantas. O estudo de campo foi instalado no delineamento em blocos casualizados. No final do estudo, aos 28 DAA, todos os tratamentos apresentaram controle total das plantas de milheto, independente da dose e do volume de aplicação, exceto para os tratamentos com paraquat + diuron e amônio-glufosinate na menor dose (25% da dose), nos dois volumes de aplicação, que apresentaram ainda controles considerados bons a muito bom, sendo que para esses dois herbicidas houve efeito do volume de calda, na qual o controle foi superior com o uso de volume menor.

The objective of this work was to evaluate the efficacy of different herbicides applied in post-emergence in the desiccation of pearl millet with different doses and spray volumes. For the desiccation, doses at 0, 25, 50 and 100% of the herbicide glyphosate (1,440 g ha-1) were used in 3 formulations (Original Round up, Round up Transorb and Round up WG), paraquat + diuron (500 + 250 g ha-1) and glufosinateammonium (800 g ha-1). In all treatments, 2 spray volumes (200 and 400 l ha-1) were used, and there was 1 treatment (control) without the application of herbicides. The treatment effects were evaluated visually at 3, 7, 14, 21 and 28 days after application (DAA). At the end of the evaluations, the dry mass of the plants was determined. The field trial was carried out in randomized block design. At the end of the study, at 28 DAA, all treatments presented total control of plant pearl millet, regardless of the dose and spray volume, except the treatments with paraquat + diuron and glufosinate-ammonium at the lower dose (25%), in 2 spray volumes, which nevertheless showed good and very good control, these two herbicides presenting an effect of volume, in which the control was higher with the use of smaller volume.

Role of hydroxyproline-rich glycoproteins in resistance of pearl millet against downy mildew pathogen Sclerospora graminicola.

Hydroxyproline-rich glycoproteins (HRGPs) are important plant cell wall components involved in plant defense response to pathogen attack. In the present study, a resistant pearl millet (Pennisetum glaucum) cultivar, IP18292, was compared with a susceptible cultivar, 7042S, to investigate the contribution of HRGPs in the successful defense against the phytopathogenic oomycete S. graminicola. Northern hybridization using MeHRGP cDNA, a heterologous probe from cassava, indicated steady accumulation of HRGP transcripts, from 2 h.p.i. onwards with a maximum at 6 h.p.i., in the resistant cultivar. This is followed by HRGPs accumulation at about 8 h.p.i. as revealed by Western-blot analysis. Immunocytochemical localization by tissue printing and confocal immunofluorescence microscopy indicated cell walls of parenchymatic cells and the vascular tissue of coleoptile as sites of HRGP deposition. In vitro studies in the presence of horseradish peroxidase and H2O2 showed cross-linking of pearl millet HRGPs, which occurred parallel to isodityrosine accumulation. Inducible high isodityrosine content was also observed in vivo in the resistant cultivar. Here, H2O2 was found to accumulate as twin burst at 1 and 6 h.p.i., whereas in the susceptible cultivar only an early single peak was detectable. Moreover, the amount of hydroxyproline in HRGPs was about twice as high in the resistant as in the susceptible cultivar. These results suggest that cell wall strengthening in S. graminicola-infected resistant pearl millet is brought about by a combination of polypeptide cross-linking of isodityrosine as well as by the high content of hydroxyproline in HRGPs, and H2O2, in contrast to the susceptible plant.

Antinutritional factor content and hydrochloric acid extractability of minerals in pearl millet cultivars as affected by germination.

Four pearl millet cultivars of two different species--Kordofani and Ugandi (Pennisetum typhoideum) and Madelkawaya and Shambat (Pennisetum glaucum)--were germinated for 6 days. The germinated grains were dried and milled. Phytic acid and polyphenol contents and hydrochloric acid (HCl) extractability of minerals from the malt flours were determined at intervals of 2 days during germination. Phytic acid and polyphenol contents decreased significantly (P <0.01) with an increase in germination time, with a concomitant increase in HCl extractable minerals. However, the major mineral content was significantly decreased while that of trace minerals was increased with germination time. When the grains were germinated for 6 days, Madelkawaya had higher extractable calcium while Ugandi had higher extractable phosphorus, whereas iron and manganese recorded high levels in Shambat and Madelkawaya, respectively. There was good correlation between antinutritional factors reduction and the increment in extractable minerals with germination time.

Molecular characterization of the genomic region linked with apomixis in Pennisetum/Cenchrus.

Apomixis is defined as asexual reproduction through seeds, although this outcome can be achieved by multiple pathways. Since little is known about the molecular control of these pathways, how they might intersect is also a mystery. Two of these pathways in the grass family, diplospory and apospory, are receiving attention from molecular biologists. Apospory in Pennisetum/Cenchrus, two genera of panicoid grasses, results in the formation of four-nucleate embryo sacs that lack antipodals. Sexual reproduction frequently aborts so that the resulting seed is composed of (1) a parthenogenetically derived embryo that is genetically identical to the mother and (2) endosperm formed through pseudogamy. The transmission of apomixis is associated with the transfer of a linkage block on a single chromosome. This linkage block contains repetitive sequences as well as hemizygous, low-copy DNA sequences. Fluorescence in situ hybridization has demonstrated that these DNA regions occur on only a single chromosome, but not its homologs, in the polyploid apomicts studied. Features of the apomixis-associated region resemble those of other chromosomal segments isolated from recombination and replete with "selfish" DNAs.