Inheritance of mitochondrial DNA in lentil (Lens culinaris Medik.)

Restriction fragment analysis was used to examine the inheritance of lentil mitochondrial DNA (mtDNA) in F₁ and F₅ progeny from intrasubspecific (Lens culinaris ssp. culinaris) crosses and in F₁ progeny from intersubspecific (Lens culinaris ssp. orientalis x L. culinaris ssp. culinaris) crosses. Southern blots of digested parental and progeny DNA were hybridized to heterologous maize mtDNA probes specific to coxI and atp6 genes. Two restriction fragment polymorphisms separated L.c. ssp. culinaris Laird and Eston from L.c. ssp. culinaris ILL5588, and one restriction fragment polymorphism distinguished L.c. ssp. culinaris Laird and Eston from L.c. ssp. orientalis LO4. Twelve of 13 f₁ progeny and all F₅ progeny from the intrasubspecific crosses, and all F₁ progeny from intersubspecific crosses had only maternal mtDNA restriction fragments. One f₁ plant from an Eston x ILL5588 cross inherited mtDNA fragments from both parents. Nuclear DNA inheritance was biparental in all F₁ progeny.NRCC No. 38451     

Agrobacterium tumefaciens-mediated beta-glucuronidase (GUS) gene expression in lentil (Lens culinaris Medik.) tissues

Summary Lentil (Lens culinaris Medik.) shoot apex, epicotyl, and root expiants were capable of expressing an intron-containing beta-glucuronidase (GUS) gene after inoculation with the disarmed Agrobacterium strain GV2260:p35SGUSINT. Expression occurred at all wound sites on these expiants except at the end of the root expiants proximal to the cotyledonary node. GUS expression was detected using both histochemical and fluorescence assays and was stable for at least nine days after inoculation for epicotyl and root expiants, and for at least seventeen days for shoot apices. Non-inoculated controls, or controls inoculated with an Agrobacterium strain lacking the GUS gene, did not produce any background blue staining in the histochemical assay. Expression levels for all lentil expiants were substantially lower than for comparable flax (Linum usitatissimum L.) expiants which served as a positive control.     

Crown gall transformation of lentil (Lens culinaris Medik.) with virulent strains of Agrobacterium tumefaciens

Summary Four diverse strains of Agrobacterium tumefaciens (C58, Ach5, GV3111, and A281) were capable of inducing tumors at a high frequency on inoculated stems of lentil (Lens culinaris Medik. cultivar Laird) in vivo, and on excised shoot apices in vitro. GV3111 and Ach5 produced the largest and heaviest tumors in vivo, while A281 produced the heaviest tumors in vitro. Tumor formation and opine production are indicative of plant cell transformation and tumors produced appropriate opines: nopaline (C58), octopine (Ach5 and GV3111), and agropine and mannopine (A281). Southern analysis of DNA from a tumor line produced by strain C58 showed that a T-DNA fragment had been transferred into the lentil genome.     

Plant regeneration of the legume Lens culinaris Medik. (lentil) in vitro

Until recently, grain legumes in general have proven recalcitrant at de novo regeneration in vitro. By culturing portions of lentil (Lens culinaris) shoot meristems and epicotyls on a medium containing kinetin and gibberellic acid, callus tissue was produced which could be induced to regenerate shoots in relatively large numbers, even after several subcultures. The shoots could be rooted in a mist chamber to yield whole, fertile plants.     

TDZ-induced direct shoot organogenesis and somatic embryogenesis on cotyledonary node explants of lentil (Lens culinaris Medik.)

An efficient and simple procedure for inducing high frequency direct shoot organogenesis and somatic embryogenesis in lentil from cotyledonary node explants (without both the cotyledons) in response to TDZ alone is reported. TDZ at concentration lower than 2.0 μM induced shoot organogenesis whereas at higher concentration (2.5–15 μM) it caused a shift in regeneration from shoot organogenesis to somatic embryogenesis. The cotyledonary node and seedling cultures developed only shoots even at high concentrations of BAP and TDZ, respectively. TDZ at 0.5 and 5.0 μM was found to be optimal for inducing an average of 4–5 shoots per cotyledonary node in 93 % of the cultures and 55 somatic embryos in 68 % of the cultures, respectively. The somatic embryos were germinated when transferred to lower TDZ concentration (0.5–1.0 μM). The shoots were rooted on MS basal medium containing 2.5 μM IBA. The plantlets were obtained within 8 weeks from initiation of culture and were morphologically similar to seed-raised plants. The possible role of stress in thidiazuron induced somatic embryogenesis is discussed.Key words: Thidiazuron, Lens culinaris, Somatic embryogenesis, Organogenesis     

Serological and molecular characteristics of pea seed borne mosaic virus-PSbMV from lentil (Lens culinaris Medik L.) in Iran

Abstract

During 2016 growing season, samples with leaf yellowing and mosaic like symptoms were collected from main lentil (Lens culinaris Medik L.) fields. Specific ELISA positive PSbMV samples were selected for RT-PCR. Using specific pair of primer towards CP gene region of the virus, PCR product of approx.235?bp was amplified. Based on the nucleotide sequences of the CP gene PSbMV isolates were classified in two major groups. In which, isolates in group I divided into two subgroups of Iranian isolates (B2) and (G1), along with Czech Republic, Australia, China, Greece, Pakistan and Egypt were placed in the subgroup-I. Isolate (V18) from Iran placed independently in group II. In the grouping based on the amino acid sequencing the isolates divided into two phylogenetic groups. Iranian isolates along with an isolate from Australia categorized in group-II, however world isolates were all clustered in group-I.     

Heat Shock-Induced Salt Stress Tolerance in Lentil (Lens culinaris Medik.)

Russian Journal of Plant Physiology - Soil salinity is a major constraint in crop production. Of the different strategies to cope with salt stress, a cross-tolerance strategy is inexpensive and...     

Effect of benzylaminopurine on in vitro and in vivo root development in lentil, Lens culinaris Medik

The effect of benzylaminopurine (BAP) on the formation of roots from lentil shoots regenerated on media containing BAP was studied. Seedling shoot tips, first nodes and bractlets, and immature seeds cultured on the initiation media containing 2.25 or 0.225 mg/l of BAP regenerated multiple bud shoots. The regenerated shoots formed roots in percentages ranging from 4.6 to 39.9% on a rooting medium (R medium) containing 2 mg/l of indoleacetic acid. Rooting success on R medium depended upon the cytokinin used in the initiation media, its concentration, and the time elapsed during shoot formation on these media prior to transplanting regenerated shoots to R medium. In vivo study of root growth of lentil seedlings demonstrated the strong inhibitory effect of BAP on root growth reflected in a drastic reduction of the mitotic index of the root meristem. Received: 27 August 1996 / Revision received: 12 December 1996 / Accepted: 15 January 1997     

Half-embryo cocultivation technique for estimating the susceptibility of pea (Pisum sativum L.) and lentil (Lens culinaris Medik.) cultivars toAgrobacterium tumefaciens

Langitudinally sliced embryonic axes from pea and lentil mature seeds cocultivated withA. tumefaciens carrying agus reporter gene in its T-DNA provided a convenient means to evaluate the efficiency of gene transfer to tissues in different cultivars and cocultivation conditions. Use of this technique demonstrated wide variation in susceptibility toAgrobacterium among several pea and lentil commercial genotypes.     

Comparative study of different cytokinins in the induction of morphogenesis in lentil (Lens culinaris Medik)

Summary This study presents the first comparative analysis of the effect of four different cytokinins, applied to different lentil (Lens culinaris Medik.) explants in four different concentrations. with regard to the regeneration of shoots and roots of the pulse crop. The variables explant, phytohormone and concentration were all found to be highly significant. Mature seed explants showed the highest shoot regeneration over all the phytohormones and concentrations tested. Thidiazuron (TDZ) and benzyladenine (BA) showed a higher number of regenerated shoots than kinetin (KIN) and zeatin (ZEA); an increase from 1.25 μM to 10 μM of any cytokinin in general doubled the number of shoots regenerated. The average length of regenerated shoots was found to be inversely proportional to the number of shoots regenerated. TDZ and BA were found to inhibit root development more than KIN and ZEA. The highest root regeneration frequency was obtained from shoots regenerated on media containing 1.25 μM ZEA. The study concludes that in order to obtain whole plants it is best to regenerate shoots on media containing the cytokinins KIN or ZEA at low concentrations, in order to be able subsequently to regenerate roots.     

GABA metabolism and ROS induction in lentil (Lens culinaris Medik) plants by synthetic 1,2,3-Thiadiazole compounds

In this study, we investigated the physiological effects of three synthetic 1,2,3-thiadiazole compounds (compound I: 4-[1,2,3]-Thiadiazole-4-yl-phenol, compound II: 1,3-Bis[4-(1,2,3-thiadiazol-4-yl)phenoxy]propane, compound III: 4-(4-{2,3,4,5,6-Penta[4-(1,2,3-thiadiazole-4-yl)phenoxymethyl]benzyloxy}phenyl)-1,2,3-thiadiazole) treatments on plants via the characterization of γ-aminobutyric acid metabolite level, the accumulation of reactive oxygen species (ROS), total protein contents, total carbohydrate contents, fresh weight and dry mass in two lentil (Lens culinaris Medik) cultivars (Jordan 1 and Jordan 2). In response to the three synthetic 1,2,3-thiadiazole compounds (I, II and III) treatments. A significant increase in γ- aminobutyric acid (GABA) and malondialdehyde (MDA) levels and a significant reduction in carbohydrates and protein levels and fresh weight and dry mass were obtained in both lentil cultivars. Jordan 2 cultivar showed the highest GABA and MDA accumulation and significant reduction in the carbohydrate and protein levels under the various thiadiazole compounds treatments. This suggests that GABA molecule may act as a defense mechanism and signaling molecule in carbohydrate metabolism and other physiological systems in lentil. In conclusion, our results revealed that the synthetic 1,2,3-thiadiazole compound ?? (1,3-Bis[4-(1,2,3-thiadiazol-4-yl)phenoxy]propane) had the strongest physiological inhibitory effect on lentil.     

Exploring genetic variability within lentil (Lens culinaris Medik.) and across related legumes using a newly developed set of microsatellite markers

Lentil (Lens culinaris Medik.) is an economically important grain legume, yet the genetic and genomic resources remain largely uncharacterized and unexploited in this crop. Microsatellites have become markers of choice for crop improvement applications. Hence, simple sequence repeat (SSR) markers were developed for lentil through the construction of genomic library enriched for GA/CT motifs. As a result 122 functional SSR primer pairs were developed from 151 microsatellite loci and validated in L. culinaris cv. Precoz. Thirty three SSR markers were utilized for the analysis of genetic relationships between cultivated and wild species of Lens and related legumes. A total of 123 alleles were amplified at 33 loci ranging from 2–5 alleles with an average of 3.73 alleles per locus. Polymorphic information content (PIC) for all the loci ranged from 0.13 to 0.99 with an average of 0.66 per locus. Varied levels of cross genera transferability were obtained ranging from 69.70 % across Pisum sativum to 12.12 % across Vigna radiata. The UPGMA based dendrogram was able to establish the uniqueness of each genotype and grouped them into two major clusters clearly resolving the genetic relationships within lentil and related species. The new set of SSR markers reported here were efficient and highly polymorphic and would add to the existing repertoire of lentil SSR markers to be utilized in molecular breeding. Moreover, the improved knowledge about intra- and inter-specific genetic relationships would facilitate germplasm utilization for lentil improvement.     

A rooting procedure for lentil (Lens culinaris Medik.) and other hypogeous legumes (pea,chickpea and Lathyrus) based on explant polarity

The present study assessed the rooting response of lentil nodal segments in relation to explant polarity, hormone, salt and carbohydrate concentrations of the medium. Nodal segments of lentil with an axillary bud cultured in an inverted orientation (apical end in medium) showed higher rooting frequencies than explants cultured in a normal orientation (basal end in medium). The highest rooting percentage (95.35%) and average number of shoots regenerated per explant (2.4) were obtained from explants placed in an inverted orientation on Murashige and Skoog (MS) medium salts with 3% sucrose, supplemented with 5 microM indole acetic acid (IAA) and 1 microM kinetin (KN). Reducing or increasing phytohormone concentration did not alter significantly root regeneration of inverted explants. Sucrose at 3% allowed higher root regeneration frequencies compared to 1.5% sucrose. MS full concentration permitted regeneration of longer shoots with more nodes per regenerated shoot, compared to MS half-strength, which regenerated more shoots of shorter length and with less nodes. Inverted nodal segments of other hypogeous legumes (pea, chickpea and Lathyrus) also exhibited higher rooting frequencies than explants cultured in a normal orientation on MS medium with 3% sucrose and supplemented with 5 microM IAA and 1 microM KN. The most novel application of this study is the culture of nodal segments of hypogeous legumes in an inverted orientation. This procedure is a considerable improvement over other published procedures concerning in vitro rooting of lentil, pea, chickpea and Lathyrus.     

Lentil (Lens culinaris Medik.) nodulates with genotypically and phenotypically diverse rhizobia in Ethiopian soils

Forty-eight lentil-nodulating rhizobia were isolated from soil samples collected from diverse agro-ecological locations in Ethiopia, and characterized based on 76 phenotypic traits. Furthermore, 26 representative strains were selected and characterized using multilocus sequence analyses (MLSA) of core (16S rRNA, recA, atpD, glnII and gyrB) and symbiotic (nodA and nifH) genes. Numerical analysis of phenotypic characteristics showed that the 48 test strains fell into three major distinct clusters. The phylogenetic trees based on 16S rRNA genes showed that they belong to the Rhizobium genus. Our phylogenetic reconstruction based on combined gene trees (recA, atpD and glnII) supported three distinct sub-lineages (Clades I–III). While genospecies I and II could be classified with Rhizobium etli and Rhizobium leguminosarum, respectively, genospecies III, might be an unnamed genospecies within the genus Rhizobium. Phylogenetic reconstruction based on the symbiosis-related genes supported a single cluster, indicating differences in the evolutionary histories between chromosomal and symbiotic genes. Overall, these results confirmed the presence of a great diversity of lentil-nodulating Rhizobium species in Ethiopia, inviting further exploration. Moreover, the differences in symbiotic effectiveness of the test strains indicated the potential for selecting and using them as inoculants to improve the productivity of lentil in the country.     

Transformation of lentil (Lens culinaris M.) cotyledonary nodes by vacuum infiltration ofAgrobacterium tumefaciens

Lentil cotyledonary nodes are some of the most regenerative tissues in legumes. Attempts to transform them by vacuum filtration have been limitedly successful. This report describes a rapid and convenient transient expression protocol based on vacuum infiltration ofAgrobacterium cells into lentil cotyledonary nodes. Vacuum-infiltrated tissues had significantly (P<.05) higher transient GUS expression than did noninfiltrated tissues. Under optimal conditions (infiltration at 200 mmHg for 20 min), 95% of theAgrobacterium-infiltrated explants exhibited an average of 16 blue foci. We believe this to be the first report of this technique for transient gene expression in lentil cotyledonary nodes.     

Genetic diversity of rhizobia nodulating lentil (Lens culinaris) in Bangladesh

In order to determine the bacterial diversity and the identity of rhizobia nodulating lentil in Bangladesh, we performed a phylogenetic analysis of housekeeping genes (16S rRNA, recA, atpD and glnII) and nodulation genes (nodC, nodD and nodA) of 36 bacterial isolates from 25 localities across the country. Maximum likelihood (ML) and Bayesian analyses based on 16S rRNA sequences showed that most of the isolates (30 out of 36) were related to Rhizobium etli and Rhizobium leguminosarum. Only these thirty isolates were able to re-nodulate lentil under laboratory conditions. The protein-coding housekeeping genes of the lentil nodulating isolates showed 89.1-94.8% genetic similarity to the corresponding genes of R. etli and R. leguminosarum. The same analyses showed that they split into three distinct phylogenetic clades. The distinctness of these clades from closely related species was also supported by high resolution ERIC-PCR fingerprinting and phenotypic characteristics such as temperature tolerance, growth on acid-alkaline media (pH 5.5-10.0) and antibiotic sensitivity. Our phylogenetic analyses based on three nodulation genes (nodA, nodC and nodD) and cross-inoculation assays confirmed that the nodulation genes are related to those of R. leguminosarum biovar viciae, but clustered in a distinct group supported by high bootstrap values. Thus, our multi-locus phylogenetic analysis, DNA fingerprinting and phenotypic characterizations suggest that at least three different clades are responsible for lentil nodulation in Bangladesh. These clades differ from the R. etli-R. leguminosarum group and may correspond to novel species in the genus Rhizobium.     

Root exudates from lentil (Lens culinaris medic) seedlings in relation to wilt disease

Summary Root exudates from healthy and diseased lentil plants (Lens culinaris) have been investigated in relation to the wilt disease caused byFusarium oxysporum f.lentis. In all ten amino acids and five sugars have been detected. The spore germination of the pathogen in root exudates indicated that 21-days root exudate was inhibitory. Glycine and phenylalanine were detected in 21-days exudate and were found to have an inhibitory effect upon the germination of the spores of the pathogen which may partly explain the lesser disease incidence when plants of more than 3 weeks are inoculated with the pathogen.     

Ribosomal and mitochondrial peroxidase isoenzymes of the lentil (Lens culinaris) root

Summary Ribosome- and mitochondria-rich preparations were obtained from lentil roots and their peroxidase isoenzymes examined by starch gel electrophoresis. Seven isoenzymes were shown to be associated with both the ribosomal and mitochondrial fractions.The apparent similarity between ribosomal and mitochondrial isoenzyme patterns, together with the observation that a considerable amount of peroxidase activity dissociated from the ribosomes during sedimentation of the ribosomal preparation into a sucrose gradient, suggested that at least some of these isoperoxidases were attached to membranes.     

Detecting DNA polymorphism and genetic diversity in Lentil (Lens culinaris Medik.) germplasm: comparison of ISSR and DAMD marker

Genetic diversity and interrelationships among 31 lentil genotypes were evaluated using 10 Inter-Simple Sequence Repeat (ISSR) and 10 directed amplification of minisatellite DNA region (DAMD) primers. A total of 43 and 48 polymorphic bands were amplified by ISSR and DAMD markers, respectively. Average polymorphism information content (PIC) for ISSR and DAMD markers were 0.37 and 0.41, respectively. All 31 lentil genotypes could be distinguished by ISSR markers into three groups and by DAMD markers into two groups. Various molecular markers show a different efficiency for evaluating DNA polymorphism in lentil and indicate that the patterns of variation are clearly influenced by the genetic marker used. Comparatively, the genetic diversity of examined lentil genotypes by two different marker techniques (ISSR and DAMD) was high and indicated that ISSR and DAMD are effective and promising marker systems for fingerprinting in lentil and give useful information on its genetic relationships.