Purification,characterization and differential hormonal regulation of a β-1,3-glucanase and two chitinases from chickpea (Cicer arietinum L.)

Chickpea (Cicer arietinum L.) cell-suspension cultures were used to isolate one -1,3-glucanase (EC 3.2.1.29) and two chitinases (EC 3.2.1.14). The -1,3-glucanase (M_r = 36 kDa) and one of the chitinases (M_r = 32 kDa) belong to class I hydrolases with basic isoelectric points (10.5 and 8.5, respectively) and were located intracellularly. The basic chitinase (BC) was also found in the culture medium. The second chitinase (M_r = 28 kDa), with an acidic isoelectric point of 5.7, showed homology to N-terminal sequences of class III chitinases and represented the main protein accumulating in the culture medium. Polyclonal antibodies raised against the basic -1,3-glucanase (BG) and the acidic chitinase (AC) were shown to be monospecific. The anti-AC antiserum failed to recognize the BC on immune blots, confirming the structural diversity between class I and class III chitinases. Neither chitinase exhibitied lysozyme activity. All hydrolases were endo in action on appropriate substrates. The BC inhibited the hyphal growth of several test fungi, whereas the AC failed to show any inhibitory activity. Expression of BG activity appeared to be regulated by auxin in the cell culture and in the intact plant. In contrast, the expression of neither chitinase was apparently influenced by auxin, indicating a differential hormonal regulation of -1,3-glucanase and chitinase activities in chickpea. After elicitation of cell cultures or infection of chickpea plants with Ascochyta rabiei, both system were found to have hydrolase patterns which were qualitatively and quantitatively comparable. Finally, resitant (ILC 3279) and susceptible (ILC 1929) cultivars of chickpea showed no appreciable differences with regard to the time and amount of hydrolase accumulation after inoculation with spores of A. rabiei.Abbreviations AC acidic chitinase - BC basic chitinase - BG = basic -1,3-glucanase - CM-Chitin-RBV carboxymethylated-chitin-remazol brilliant violet - 2,4-D 2,4-dichlorophenoxyacetic acid - ILC international legume chickpea - M_r relative molecular mass - pI isoelectric point - SDS-PAGE sodium dodecyl sulfatepolyacrylamide gel electrophoresis We thank the Deutsche Forschungsgemeinschaft and Fonds der Chemischen Industrie for financial support and ICARDA, Aleppo, Syria, for the provision of seed material. We also thank Dr. B. Fritig (Institut de Biologie Moléculaire des Plantes, CNRS, Straßbourg, France) and Dr. F. Meins, Jr. (Friedrich-Miescher-Institut, Basel, Switzerland) for their kind gifts of antibodies.     

A linkage map of the chickpea (Cicer arietinum L.) genome based on recombinant inbred lines from a C. arietinum×C. reticulatum cross: localization of resistance genes for fusarium wilt races 4 and 5

An integrated molecular marker map of the chickpea genome was established using 130 recombinant inbred lines from a wide cross between a cultivar resistant to fusarium wilt caused by Fusarium oxysporum Schlecht. emend. Snyd. &. Hans f. sp. ciceri (Padwick) Snyd & Hans, and an accession of Cicer reticulatum (PI 489777), the wild progenitor of chickpea. A total of 354 markers were mapped on the RILs including 118 STMSs, 96 DAFs, 70 AFLPs, 37 ISSRs, 17 RAPDs, eight isozymes, three cDNAs, two SCARs and three loci that confer resistance against different races of fusarium wilt. At a LOD-score of 4.0, 303 markers cover 2077.9 cM in eight large and eight small linkage groups at an average distance of 6.8 cM between markers. Fifty one markers (14.4%) were unlinked. A clustering of markers in central regions of linkage groups was observed. Markers of the same class, except for ISSR and RAPD markers, tended to generate subclusters. Also, genes for resistance to races 4 and 5 of fusarium wilt map to the same linkage group that includes an STMS and a SCAR marker previously shown to be linked to fusarium wilt race 1, indicating a clustering of several fusarium-wilt resistance genes around this locus. Significant deviation from the expected 1 : 1 segregation ratio was observed for 136 markers (38.4%, P<0.05). Segregation was biased towards the wild progenitor in 68% of the cases. Segregation distortion was similar for all marker types except for ISSRs that showed only 28.5% aberrant segregation. The map is the most extended genetic map of chickpea currently available. It may serve as a basis for marker-assisted selection and map-based cloning of fusarium wilt resistance genes and other agronomically important genes in future. Received: 17 November 1999 / Accepted: 4 June 2000     

Three genetic systems controlling growth, development and productivity of rice (Oryza sativa L.): a reevaluation of the ‘Green Revolution’

The Green Revolution (GR-I) included worldwide adoption of semi-dwarf rice cultivars (SRCs) with mutant alleles at GA20ox2 or SD1 encoding gibberellin 20-oxidase. Two series of experiments were conducted to characterize the pleiotropic effects of SD1 and its relationships with large numbers of QTLs affecting rice growth, development and productivity. The pleiotropic effects of SD1 in the IR64 genetic background for increased height, root length/mass and grain weight, and for reduced spikelet fertility and delayed heading were first demonstrated using large populations derived from near isogenic IR64 lines of SD1. In the second set of experiments, QTLs controlling nine growth and yield traits were characterized using a new molecular quantitative genetics model and the phenotypic data of the well-known IR64/Azucena DH population evaluated across 11 environments, which revealed three genetic systems: the SD1-mediated, SD1-repressed and SD1-independent pathways that control rice growth, development and productivity. The SD1-mediated system comprised 43 functional genetic units (FGUs) controlled by GA. The SD1-repressed system was the alternative one comprising 38 FGUs that were only expressed in the mutant sd1 backgrounds. The SD1-independent one comprised 64 FGUs that were independent of SD1. GR-I resulted from the overall differences between the former two systems in the three aspects: (1) trait/environment-specific contributions; (2) distribution of favorable alleles for increased productivity in the parents; and (3) different responses to (fertilizer) inputs. Our results suggest that at 71.4 % of the detected loci, a QTL resulted from the difference between a functional allele and a loss-of-function mutant, whereas at the remaining 28.6 % of loci, from two functional alleles with differentiated effects. Our results suggest two general strategies to achieve GR-II (1) by further exploiting the genetic potential of the SD1-repressed and SD1-independent pathways and (2) by restoring the SD1-mediated pathways, or ‘back to the nature’ to fully exploit the genetic diversity of those loci in the SD1-mediated pathways which are virtually inaccessible to most rice-breeding programs worldwide that are exclusively based on sd1.     

Organogenesis from ‘Passe Crassane’ and ‘Old Home’ pear (Pyrus communis L.) protoplasts and isoenzymatic trueness-to-type of the regenerated plants

Summary Large numbers of highly viable mesophyll protoplasts were isolated from shoot cultures of the scion cv Passe Crassane and the rootstock genotype Old Home of common pear (Pyrus communis L.). Protoplasts were cultured for both genotypes either as liquid layers or as liquid-over-agar cultures, in ammonium-free MS medium with 0.5 M mannitol, 50 mg/l casein enzymatic hydrolysate (CEH), 2.0 mg/l NAA and 1.0 mg/l BAP, plus either 0.5 mg/l IAA (for Old Home) or 2.0 mg/l IAA (for Passe Crassane). Protoplast microcalli, obtained by day 60 (Passe Crassane) or day 80 (Old Home), were transferred for further growth to ammonium-free MS medium with 2.0 mg/l NAA and 1.0 mg/l BAP. Shoot bud regeneration from the protoplastderived callus was first attempted between 100 (Passe Crassane) and 120 (Old Home) days after protoplast isolation. For Passe Crassane, shoot buds were regenerated (day 130) on a half-strength MS medium with 0.1 mg/l IBA, 0.5 mg/l BAP, 50 mg/l CEH and 20 mg/l Ca-panthotenate. For Old Home, shoot but regeneration only occurred 30 days later and on the same medium as above, which was additionally supplemented with double the concentration of the group B vitamins found in the original MS formulation and 0.05 mg/l GA₃. Following micropropagation and in vitro rooting of shoots, the plants were transferred to soil following standard procedures. Trueness-to-type of the regenerated plants was assessed by analysing their leaf isozyme banding profiles (for EST, AP, PRX, SOD, ENP, LAP, PGI, AAT, ADH, MDH and PGM) and comparing them to those corresponding to the original shoots that provided the protoplasts. No differences between the mother shoots and the protoclones were observed for any one of the 11 isozyme systems studied.     

A genetic map of soybean (Glycine max L.) using an intraspecific cross of two cultivars: ‘Minosy’ and ‘Noir 1’

Genetic markers were mapped in segregating progeny from a cross between two soybean (Glycine max (L.) Merr.) cultivars: Minsoy (PI 27.890) and Noir 1 (PI 290.136). A genetic linkage map was constructed (LOD 3), consisting of 132 RFLP, isozyme, morphological, and biochemical markers. The map defined 1550cM of the soybean genome comprising 31 linkage groups. An additional 24 polymorphic markers remained unlinked. A family of RFLP markers, identified by a single probe (hybridizing to an interspersed repeated DNA sequence), extended the map, linking other markers and defining regions for which other markers were not available.     

Effects of the planting density on water relations and production of ‘Chemlali’ olive trees (Olea europaea L.)

Water relations are a key factor limiting olive production. In this study, effects of plating density on physiological aspects and productivity of ‘Chemlali’ olive trees were analyzed under rain-fed conditions in four planting densities (156, 100, 69 and 51 trees ha⁻¹), in an experimental olive orchard located in the center of Tunisia. Seasonal changes in leaf relative water content (RWC), leaf water potential, stomatal conductance (g _s), CO₂ assimilation rate and tree production were studied. Accompanying the changes in leaf water status, all the monitored trees reduced leaf stomatal conductance (g _s) and photosynthetic rate (A) throughout the summer drought, mirroring the increase in soil moisture deficit and vapor pressure deficit. However, the decrease in gas exchange was much more pronounced in high planting densities than in low ones. Our results confirm that the increase of tree-to-tree water competition with planting density was significant in the dry climate of Tunisia. Thus, planting density is critical when planting new olive orchards in arid regions.     

Nucleotide sequence of cDNA clones encoding the complete ‘23 kDa’ and ‘16 kDa’ precursor proteins associated with the photosynthetic oxygen-evolving complex from spinach

We present the nucleotide sequences and derived amino acid sequences of cDNAs that encode the complete precursors of the extrinsic ‘23 kDa’ and ‘16 kDa’ polypeptides associated with the photosynthetic oxygen-evolving complex from spinach. The luminal proteins consist of 267/186 (precursor/mature 23 kDa protein) and 232/149 (16 kDa polypeptide) amino acid residues corresponding to molecular masses of 28.5/20.2 and 24.9/16.5 kDa, respectively. Secondary structure predictions disclose epitopes that are potential candidates for two-step processing of the precursors during import and intraorganelle routing as well as for calcium sequestering, chloride binding and subunit/subunit interaction.     

QTL analysis of resistance to sharka disease in the apricot (Prunus armeniaca L.) ‘Polonais’ × ‘Stark Early Orange’ F1 progeny

Different hypotheses on the genetic control of the resistance to the plum pox virus (PPV) have been reported in apricot, but there was a lack of agreement about the number of loci involved. In recent years, apricot genetic maps have been constructed from progenies derived from ‘Stark Early Orange’ or ‘Goldrich’, two main sources of resistance, three of these including the mapping of the PPV resistance loci. As the location of the locus was not precisely established, we mapped the PPV resistance loci using interval mapping (IM), composite interval mapping (CIM), and the Kruskal–Wallis non-parametric test in the F₁ progeny derived from a cross between the susceptible cv. ‘Polonais’ and ’Stark Early Orange’. Four genomic regions were identified as being involved in PPV resistance. One of these mapped to the upper region of linkage group 1 of ‘Stark Early Orange’, and accounted for 56% of the phenotypic variation. Its location was similar to the one previously identified in ‘Goldrich’ and Prunus davidiana. In addition, a gene strongly associated to these major quantitative trait loci (QTL) was found to be related to PPV infection. Two putative QTLs were detected on linkage groups 3 of ‘Polonais’ and 5 of both ‘Polonais’ and ‘Stark Early Orange’ with both parametric and non-parametric methods at logarithm of odds (LOD) scores slightly above the detection threshold. The last QTL was only detected in the early stage of the infection. PPV resistance is, thus, controlled by a major dominant factor located on linkage group 1. The hypothesis of recessive factors with lower effect is discussed.     

Monosomic analysis of heading date and spikelet number in the common wheat (Triticum aestivum L.) multispikelet line ‘Noa’

Summary Genetic analysis of heading date and spikelet number was carried out in the common wheat (Triticum aestivum L.) multispikelet line Noa, by using the monosomic series of the regular line Mara. Noa's high number of spikelets was found to be controlled by a recessive major gene on chromosome 2D; a slight reduction in spikelet number was induced by another recessive gene on Noa's 7A chromosome. Noa's late heading date was found to be controlled by two recessive genes, located on chromosome 2D (a major effect) and 6B (a minor effect). The nature of the genes located on Noa's 2D chromosome and the relationship between spikelet number and heading date are discussed.     

The effects of external potassium and magnesium concentrations on the magnesium and potassium inflow rates and growth of micropropagated cherry rootstocks, “F.12/1” (Prunus avium L.) and “Colt” (Prunus avium L.) × Prunus pseudocerasus L.)

The effects (and interaction) of two solution concentrations of Mg (50, 500, μM) and two of K (250, 4250 μM) on the growth of micropropagated plants of “F. 12/1” and “Colt” were investigated using a flowing solution culture system. Magnesium inflow and growth of “Colt” and “F. 12/1” were inhibited to a similar extent by an increased concentration of K in the nutrient solution. However, the consequences of this inhibition were different. Reduced inflow of Mg in “F. 12/1” caused Mg deficiency symptoms at high and low concentrations of K, whereas this only occurred with a combination of high K concentration and low Mg concentration in “Colt”. The distribution of dry matter within the plant was significant in determining susceptibility to Mg deficiency. Since “F. 12/1” has a smaller root:shoot ratio than Colt it is unable to sustain the same concentration of Mg in leaves as “Colt” irrespective of external K concentration. The molar ratio of K:Mg in soil solutions should remain <8.5:1 in order to ensure maximum growth of “F. 12/1” and “Colt”. This revised version was published online in June 2006 with corrections to the Cover Date.     

The chemical compound ‘Heatin’ stimulates hypocotyl elongation and interferes with the Arabidopsis NIT1-subfamily of nitrilases

Temperature passively affects biological processes involved in plant growth. Therefore, it is challenging to study the dedicated temperature signalling pathways that orchestrate thermomorphogenesis, a suite of elongation growth-based adaptations that enhance leaf-cooling capacity. We screened a chemical library for compounds that restored hypocotyl elongation in the pif4-2–deficient mutant background at warm temperature conditions in Arabidopsis thaliana to identify modulators of thermomorphogenesis. The small aromatic compound ‘Heatin’, containing 1-iminomethyl-2-naphthol as a pharmacophore, was selected as an enhancer of elongation growth. We show that ARABIDOPSIS ALDEHYDE OXIDASES redundantly contribute to Heatin-mediated hypocotyl elongation. Following a chemical proteomics approach, the members of the NITRILASE1-subfamily of auxin biosynthesis enzymes were identified among the molecular targets of Heatin. Our data reveal that nitrilases are involved in promotion of hypocotyl elongation in response to high temperature and Heatin-mediated hypocotyl elongation requires the NITRILASE1-subfamily members, NIT1 and NIT2. Heatin inhibits NIT1-subfamily enzymatic activity in vitro and the application of Heatin accordingly results in the accumulation of NIT1-subfamily substrate indole-3-acetonitrile in vivo. However, levels of the NIT1-subfamily product, bioactive auxin (indole-3-acetic acid), were also significantly increased. It is likely that the stimulation of hypocotyl elongation by Heatin might be independent of its observed interaction with NITRILASE1-subfamily members. However, nitrilases may contribute to the Heatin response by stimulating indole-3-acetic acid biosynthesis in an indirect way. Heatin and its functional analogues present novel chemical entities for studying auxin biology.     

Antipyretic activity of Caesalpinia digyna (Rottl.) leaves extract along with phytoconstituent’s binding affinity to COX-1, COX-2, and mPGES-1 receptors: In vivo and in silico approaches

Caesalpinia digyna (Rottl.) (Family: Fabaceae) is well known for its numerous medicinal values against several human disorders including fever, senile pruritis, diarrhea, tuberculosis, tonic disorder, diabetes, etc. The current study is intended to investigate the in vivo antipyretic activity of the methanol extract of C. digyna leaves (MECD) and its carbon-tetrachloride (CTCD) and butanol fraction (BTCD). Besides, in silico molecular docking and ADME/T profiling of the selective identified bioactive compounds of C. digyna has been also studied to validate the experimental outcomes and establish a better insight into the possible receptor-ligand interaction affinity. In vivo antipyretic activity of MECD, CTCD and BTCD were evaluated by employing yeast induced pyrexia technique in mice model and in silico analysis of the identified compounds of C. digyna has been implemented using PyRx autodock vina, Discovery Studio 2020, UCSF Chimera software and ADME/T online tools. MECD and BTCD unveiled significant antipyretic activity in dose dependent manner whereas, CTCD failed to exhibit significant antipyretic activity. Comparing to other test sample, MECD (400 mg/kg; b.w) (p < 0.001) displayed maximum inhibition of pyrexia. In molecular docking approach, docking score between −6.60 to −10.20 kcal/mol have been revealed. Besides, in ADME/T analysis, no compound violated the lipiniski’s 5 rules and displayed any toxicity. Biological and computational approaches ascertain the ethno-botanical use of C. digyna as a good agent against pyrexia and the compounds of C. digyna are primarily proved as safe. Hereafter, further analysis is suggested to validate this research.     

A ‘Chinese Spring’ wheat (Triticum aestivum L.) bacterial artificial chromosome library and its use in the isolation of SSR markers for targeted genome regions

A bacterial artificial chromosome (BAC) library was constructed from the bread wheat (Triticum aestivum L.) genotype ‘Chinese Spring’ (‘CS’). The library consists of 395,136 clones with an estimated average insert size of 157 kb. This library provides an estimated 3.4-fold genome coverage for this hexaploid species. The genome coverage was confirmed by RFLP analysis of single-copy RFLP clones. The CS BAC library was used to develop simple sequence repeat (SSR) markers for targeted genome regions using five sequence-tagged-site (STS) markers designed from the chromosome arm of 3BS. The SSR markers for the targeted genome region were successfully obtained. However, similar numbers of new SSR markers were also generated for the other two homoeologous group 3 chromosomes. This data suggests that BAC clones belonging to all three chromosomes of homoeologous group 3 were isolated using the five STS primers. The potential impacts of these results on marker isolation in wheat and on library screening in general are discussed.     

The effect of total inorganic nitrogen and the balance between its ionic forms on adventitious bud formation and callus growth of ‘Węgierka Zwykła’ plum (<Emphasis Type="Italic">Prunus domestica</Emphasis> L.)

The effect of different ammonium NH ₄ ⁺ and nitrate NO ₃ ^? ratios (4:1, 2:1, 1:1, 1:2, 1:4, 1:6) on organogenesis of ‘W?gierka Zwyk?a’ leaf explants cultivated on media with nitrogen levels equalling full- or half-MS was investigated. On media with total nitrogen equal to ½ MS, explant regeneration increased significantly and was highest on media with 1:2 or 1:4 NH ₄ ⁺ :NO ₃ ^? ratio. An excess of ammonium versus nitrate ions had a negative effect on both regeneration and biomass. Addition of potassium to the medium increased the fresh weight of explants and the number of adventitious buds.     

Pest control in museums: toxicity of para-dichlorobenzene, ‘Vapona’™, and naphthalene against all stages in the life-cycle of museum pests,Dermestes maculatus Degeer,and Anthrenus verbasci (L.) (Coleoptera: Dermestidae)

Museums, herbaria, libraries and archival collections have traditionally relied on chemicals for the prevention and treatment of pest infestations. While current evidence suggests that the use of chemicals is declining, however, they are still found in many collections. The efficacy of three ‘insecticides’, para-dichlorobenzene, ‘Vapona’ and naphthalene (used in some museums to treat localized infestations and for their apparent residual benefit) against two insect species (Coleoptera: Dermestidae) was evaluated. Despite considerable variation in insect susceptibility, ‘Vapona’ was found in general, to be the most effective of the three chemicals used, particularly against larvae and adults. Naphthalene was the least effective, with low mortality rates recorded in the majority of the insect stages tested. Based on this study, an exposure/mortality relationship is presented for the prevention and treatment of insect infestations in museum and archival collections.