Isozyme and cytological markers of some Psathyrostachys juncea accessions

Summary Psathyrostachys juncea (synonymous to Elymus junceus; 2n=2x=14, NN) has unique biotic and abiotic attributes that could contribute towards wheat improvement. The effectiveness of such an intergeneric hybridization program depends greatly on being able to establish diagnostic markers of the alien chromosomes. Isoelectric focusing (IEF) analyses of six enzyme systems have identified five biochemical markers — malate dehydrogenase (MDH), esterase (EST), shikimate dehydrogenase (SKDH), phosphoglucomutase (PGM), and -amylase (-AMY) — to be of positive diagnostic value; glucosephosphate isomerase (GPI) banding profiles were of no definite value in the background of Triticum aestivum cvs Chinese Spring and Seri-82, the potential recipients of Ps. juncea chromosomes. The Giemsa C-banding karyotype distinctively separates the Ps. Juncea chromosomes from each other and from those of T. aestivum with little banding site polymorphisms prevalent among its accessions analyzed, indicating the usefulness of C-bands as cytological markers.     

Hybridization between Triticum aestivum L. and Agropyron michnoi Roshev.

Summary Intergeneric hybrids between Triticum aestivum cv Chinese Spring (2n=6x=42, AABBDD) and Agropyron michnoi Roshev. (2n=4x=28, PPPP) were obtained by embryo culture. Their spike characteristics were similar to those of common wheat but, unlike their parents, they were long-awned. The average meiotic chromosome pairing at MI of F₁ hybrids was: 6.39 I +3.75 rodII+8.64 ringII+0.81 III+0.30 IV+0.04 V, the bivalent and multivalent formation of which was much higher than expected from the genomic formulae. It is especially worthwhile to note that the F₁ hybrids were self-fertile, self set being 0.15%, and seeds were easily obtained from the backcross of f₁ plants with hexaploid and tetraploid wheats; here the seed set was more than 20.0%. The polyploid taxa and the position of A. Michnoi in Agropyron are discussed.     

Production and cytogenetics of Triticum aestivum L. hybrids with some rhizomatous Agropyron species

Summary Intergeneric hybrids between Triticum aestivum L. and conventional rhizomatous Agropyron species were produced in variable frequencies. They were recovered in high percentage frequencies for T. aestivum cultivars with A. acutum (14.6%), A. intermedium (48.0%), A. pulcherrimum (53.3%), and A. trichophorum (46.6%). The crossability percentages with the highly crossable cultivar Chinese Spring for these Agropyron species accessions were 33.12%, 65.0%, 53.3%, and 65.4%, respectively. Autosyndetic associations of two of their three genomes gave mean meiotic chromosome association data of 17.0 I (univalents) +1.53 II (ring bivalents) + 7.04 II (rod bivalents) +1.43 III (trivalents) +0.05 IV (quadrivalents) +0.01 IV (pentavalents) for A. acutum and of 21.8 I + 1.56 II (rings) +7.22 II (rods) +0.84 III + 0.04 IV for A. intermedium. Chromosome pairing at metaphase I was comparatively lower for A. pulcherrimum (34.4 I + 0.2 II (rings) +3.4 II (rods) +0.14 III) and A. trichophorum (36.7 I + 0.35 II (rings) +2.26 II (rods) + 0.04 III) hybrids with T. aestivum. Hybrids of wheat with A. campestre and A. repens were obtained in low frequency. Direct crossing did not permit T. aestivum/ A. desertorum hybridization. However, by utilizing the 2n=10x=70 A. repens/A. desertorum amphiploid as the pollen source, hybridization with T. aestivum did indeed occur. Aneuploidy was prevalent in this hybrid combination while all other hybrid combinations were apparently normal.     

Production and cytogenetic analysis of BC1, BC2, and BC3 progenies of an intergeneric hybrid between Triticum aestivum (L.) Thell. and tetraploid Agropyron cristatum (L.) Gaertn.

Summary Intergeneric hybrids between Triticum aestivum cv Chinese Spring and Agropyron cristatum 4x (2n= 5x=35, ABDPP genomes) with a high level of homoeologous meiotic pairing between the wheat chromosomes were backcrossed 3 times to wheat. Pollination of the F₁ hybrid with Chinese Spring resulted in 22 BC₁ seeds with an average seed set of 1.52%. Five BC₁ plants with 39–41 chromosomes were raised using embryo rescue techniques. Chromosome pairing in the BC₁ was characterized by a high frequency of multivalent associations, but in spite of this there was no evidence of homoeologous pairing between chromosomes of wheat and those of Agropyron. All of the plants were self sterile. The embryo rescue technique was again essential to produce 39 BC₂ plants with chromosome numbers ranging from 37 to 67. The phenomenon of meiotic non-reduction was also observed in the BC₃ progenies. In this generation male and female fertility greatly increased, and meiotic pairing was fairly regular. Some monosomic (2n=43) and double monosomic (2n=44) lines were produced. Analysis of these progenies should permit the extraction of the seven possible wheat-Agropyron disomic addition lines including those with the added chromosomes carrying the genes involved in meiotic non-reduction and in suppression of Ph activity.     

RELP markers linked to two Hessian fly-resistance genes in wheat (Triticum aestivum L.) from Triticum tauschii (coss.) Schmal

Summary Restriction fragment length polymorphism (RFLP) markers linked to genes controlling Hessian fly resistance from Triticum tauschii (Coss.) Schmal. were identified for two wheat (Triticum aestivum L.) germ plasm lines KS89WGRC3 (C3) and KS89WGRC6 (C6). Forty-six clones with loci on chromosomes of homoeologous group 3 and 28 clones on those of group 6 were surveyed for polymorphisms. Eleven and 12 clones detected T. tauschii loci in the two lines, respectively. Analysis of F₂ progenies indicated that the Hessian fly resistance gene H23 identified in C3 is linked to XksuH4 (6.9 cM) and XksuG48 (A) (15.6 cM), located on 6D. The resistance gene H24 in C6 is linked to XcnlBCD451 (5.9 cM), XcnlCD0482 (5.9 cM) and XksuG48 (B) (12.9 cM), located on 3DL.Paper No. 810 of the Cornell Plant Breeding Series     

Characterization of Thinopyrum bessarabicum chromosome segments in wheat using random amplified polymorphic DNAs (RAPDs) and genomic in situ hybridization

Ten random amplified polymorphic DNA (RAPD) markers specific to chromosome 5E^b of Thinopyrum bessarabicum were detected. Genomic in situ hybridization and standard cytological observations revealed that six of the markers are located on the 5E^b short arm and four are located on the 5E^b long arm. These RAPD markers have been used to confirm the identity of putative 5E^b (5A) and 5E^b (5D) substitution individuals. The potential of RAPDs for the detection of wheat/alien recombinants is discussed.     

Extent of genetic variability of endosperm esterases in Triticum aestivum L. 2n=6x=42

Summary Genetic variability of endosperm esterase has been studied in 42 cultivars of Triticum aestivum L. 2n=6x=42. Different techniques, including sequential electrophoresis and electrofocusing, have been used with various substrates and esterase inhibitors. The electrophoretic patterns in each cultivar are described. Chromosomal location using the nullitetrasomic and ditelosomic lines of Chinese Spring was carried out in order to relate and/or locate the esterase genes to specific chromosomes. Most of the esterase isozymes located were in the long arm of the chromosomes of the homoeology group 3; but we have found six located in the short arms, five of them in the chromosome 3AS and one in the 3DS. This location increases the number of esterase genes described, because no esterase genes had been described so far in short arms of chromosomes of the homoeology group 3. The genetic control is discussed and, according to our results, between 12 and 15 loci, organized in five compound loci, control the endosperm esterases in wheat. Also one modifier gene modifying the mobility of two esterase bands and present in all the cultivars studied is postulated.This work was supported by a personal grant (L. Rebordinos) from the P.F.P.I. and by an institutional grant from the C.A.I.C.Y.T. (PB85-0153)     

Isolation and identification of Triticum aestivum L. em. Thell. cv Chinese Spring-T. peregrinum Hackel disomic chromosome addition lines

Analyses of RFLPs, isozymes, morphological markers and chromosome pairing were used to isolate 12 Triticum aestivum cv Chinese Spring (genomes A, B, and D)-T. peregrinum (genomes S^v and U^v) disomic chromosome addition lines. The evidence obtained indicates that each of the 12 lines contains an intact pair of T. peregrinum chromosomes. One monosomic addition line, believed to contain an intact 6S^v chromosome, was also isolated. A CS-7U^v chromosome addition line was not obtained. Syntenic relationships in common with the standard Triticeae arrangement were found for five of the seven S^v genome chromosomes. The exceptions were 4S^v and 7S^v. A reciprocal translocation exists between 4S¹ and 7S¹ in T. longissimum and evidence was obtained that the same translocation exists in T. peregrinum. In contrast, evidence for syntenic relationships in common with the standard Triticeae arrangements were found for only one U^v chromosome of T. peregrinum.; namely, chromosome 2U^v. All other U^v genome chromosomes are involved in at least one translocation, and the same translocations were found in the U genome of T. umbellulatum. Evidence was also obtained indicating that the centromeric regions of 4U and 4U^v are homoeologous to the centromeric regions of Triticeae homoeologous group-6 chromosomes, that the centromeric regions of 6U and 6U^v are homoeologous to the centromeric regions of group-4 chromosomes, and that 4U and 4U^v are more closely related overall to Triticeae homoeologous group-6 chromosomes than they are to group-4 chromosomes.     

Production,morphology, and cytogenetic analysis of Elymus caninus (Agropyron caninum) x Triticum aestivum F1 hybrids and backcross-1 derivatives

Summary Intergeneric hybrids were produced between common wheat, Triticum aestivum (2n=6x=42, AABBDD) and wheatgrass, Etymus caninus (Agropyron caninum) (2n=4x=28, SSHH) — the first successful report of this cross. Reciprocal crosses and genotypes differed for percent seed set, seed development and F₁ hybrid plant production. With E. caninus as the pollen parent, there was no hybrid seed set. In the reciprocal cross, seed set was 23.1–25.4% depending upon wheat genotype used. Hybrid plants were produced only by rescuing embryos 12–13 days post pollination with cv Chinese Spring as the wheat parent. Kinetin in the medium facilitated embryo germination but inhibited root development and seedling growth. The hybrids were vigorous, self sterile, and intermediate between parents. These had expected chromosome number (2n=5x=35, ABDSH), very little chromosome pairing (0.51 II, 0.04 III) and some secondary associations. The hybrids were successfully backcrossed with wheat. Chromosome number in the BC₁ derivatives varied 54–58 with 56 as the modal class. The BC₁ derivatives showed unusually high number of rod bivalents or reduced pairing of wheat homologues. These were sterile and BC₂ seed was produced using wheat pollen.     

The chromosomal location of factors determining the presence of phenolic compounds in wheat (Triticum aestivum L.)

Summary Using thin-layer chromatography and nulli-tetrasomic and ditellosomic series of Triticum aestivum L. cv. Chinese Spring, it has been possible to relate the phenolic compounds found in adult plant leaves and 12 day-old seedling leaves with the chromosomes or chromosome arms 1 B, 2 BL, 3 BL, 5 A, 6 AL, 7 B and 7 DS.     

Isolation and analysis of endophytic microorganisms in wheat (Triticum aestivum L.) leaves

The present investigation was undertaken in order to select the surface-sterilization technique most efficient for eliminating epiphytes, to document the spectrum of endophytes of healthy leaves from three wheat cultivars in Buenos Aires Province (Argentina) and to determine their infection frequencies at three growth stages. Surface-sterilization with undiluted commercial solution of sodium hypochlorite was reaffirmed as adequate for removing epiphytes on wheat leaves. From the 450 wheat leaf segments incubated, three bacterial isolates and 130 fungal isolates were obtained. From all the isolates, 19 fungal species were identified. Bacterial isolates were characterized as Bacillus sp. There were significant differences between microorganisms, stages of growth, and stages × microorganisms interaction. Differences between cultivars, stages × cultivars, microorganisms × cultivars and for the triple interaction were not significant. Frequency of microorganisms isolated increased with crop age, but it was statistically similar for the three wheat cultivars tested (Klein Centauro, Klein Dragón and Buck Ombú). Rhodotorula rubra, Alternaria alternata, Cladosporium herbarum and Epicoccum nigrum were isolated in the highest frequency. The other microorganisms were present at intermediate or low values. The species isolated may be assigned to three groups: (a) well-known and economically important pathogens of wheat, (b) commonly abundant phylloplane fungi considered to be primary saprobic and minor pathogens and (c) species occasionally present in wheat.     

Accelerated production of transgenic wheat (Triticum aestivum L.) plants

We have developed a method for the accelerated production of fertile transgenic wheat (Triticum aestivum L.) that yields rooted plants ready for transfer to soil in 8–9 weeks (56–66 days) after the initiation of cultures. This was made possible by improvements in the procedures used for culture, bombardment, and selection. Cultured immature embryos were given a 4–6 h pre-and 16 h post-bombardment osmotic treatment. The most consistent and satisfactory results were obtained with 30 g of gold particles/bombardment. No clear correlation was found between the frequencies of transient expression and stable transformation. The highest rates of regeneration and transformation were obtained when callus formation after bombardment was limited to two weeks in the dark, with or without selection, followed by selection during regeneration under light. Selection with bialaphos, and not phosphinothricin, yielded more vigorously growing transformed plantlets. The elongation of dark green plantlets in the presence of 4–5 mg/l bialaphos was found to be reliable for identifying transformed plants. Eighty independent transgenic wheat lines were produced in this study. Under optimum conditions, 32 transformed wheat plants were obtained from 2100 immature embryos in 56–66 days, making it possible to obtain R3 homozygous plants in less than a year.     

The effect of polyamines on protein kinase activities of wheat (Triticum aestivum) L. anthers

Protein kinase activities were extracted from anther tissue of wheat (Triticum aestivum L. cv. Zaragoza) and characterized with regard to the effects of polyamines, second messengers and plant growth hormones. The protein kinases were inhibited by polyamines and cyclic nucleotides, but were stimulated by the addition of auxins, gibberellic acid and kinetin. The dominant polyamine-sensitive kinase activity was partially purified and characterized. The optimal pH of the reaction was 7.5 to 8.0 and casein was the preferred exogenous substrate. Polyamines were inhibiting in the decreasing order of putrecine > spermidine > spermine. The results are discussed against the context of the anther culture technique.     

Cytological analysis on the distribution and origin of the alien chromosome pair conferring blue aleurone color in several European common wheat (Triticum aestivum L.) strains

Summary Meiotic chromosome pairing and Giemsa C-banding analyses in crosses of several European blue-grained wheat strains with Chinese Spring double ditelosomic and other aneuploid lines showed that Triticum aestivum Blaukorn strains Berlin, Probstdorf, Tschermak, and Weihenstephan are chromosome substitutions, in which the complete wheat chromosome 4A pair is replaced, whereas the strains Brünn and Moskau are 4B substitutions. The alien chromosome pair in all of these strains is an A genome chromosome (4A) from diploid Triticum monococcum or T. boeoticum not present in common tetraploid and hexaploid cultivated wheats. The Blaukorn strain Weihenstephan W 70a86 possesses, in addition to a rye chromosome pair 5R compensating for the loss of part of chromosome 5D, a 4A/5DL translocation replacing chromosome pair 4B of wheat.     

Identification of C-banded chromosomes in meiosis of common wheat,Triticum aestivum L.

Summary The C-banding pattern of nine meiotic chromosomes of common wheat (Triticum aestivum L.) as described. In F₁s of crosses between monosomics of Chinese Spring and two Spanish wheat cultivars, univalent chromosomes were used to aid the recognition and analysis of the C-banding pattern for the individual chromosomes. The identification of one chromosome involved in one translocation in Chinese Spring x Pané 247 has been made through heterochromatin bands observed in the chromosomes involved in multivalents.     

The cytogenetics of a Triticum turgidum x Psathyrostachys juncea hybrid and its backcross derivatives

Psathyrostachys juncea (2n = 2x = 14, NN), a source of barley yellow dwarf (BYDV) virus resistance with tolerance to drought and salinity, has been successfully hybridized in its autotetraploid form (2n = 4x = 28, NNNN) as the pollen parent to durum wheat (Triticum turgidum L.). The 2n = 4x = 28 (ABNN) F₁ hybrid has a mean meiotic metaphase-I configuration of 20.29 univalents + 0.29 ring bivalents + 3.36 rod bivalents + 0.14 trivalents. Spike length, internode length, glume awn length and lemma awn length, as well as the general spike morphology of the F₁ hybrid, are intermediate with those of the two parents. Pollinating the ABNN F₁ hybrid has given backcross (BC)-I derivatives of an amphiploid (AABBNN) that expresses limited self-fertility. BC-2 derivatives have been obtained from these plants. Direct transfers of useful genes from Ps. juncea to wheat would require substantial genetic manipulation strategies. Both conventional and novel methodologies, which may complement each other, and so facilitate reaching an agricultural objective end point, are addressed.     

Enzyme activities associated with developing wheat(Triticum aestivum L.) grain amyloplasts

Intact amyloplasts from endosperm of developing wheat grains have been isolated by first preparing the protoplasts and then fractionating the lysate of the protoplasts on percoll and ficoll gradients, respectively. Amyloplasts isolated as above were functional and not contaminated by cytosol or by organelles likely to be involved in carbohydrate metabolism. The enzyme distribution studies indicated that ADP-glucose pyrophosphorylase and starch synthase were confined to amyloplasts, whereas invertase, sucrose synthase, UDP-glucose pyrophosphorylase, hexokinase, phosphofructokinase-2 and fructose-2,6-P₂ase were absent fro the amyloplast and mainly confined to the cytosol. Triose-P isomerase, glyceraldehyde-3-P dehydrogenase, phosphohexose isomerase, phosphoglucomutase, phosphofructokinase, aldolase, PPi-fructose-6-P-1 phosphotransferase, and fructose-l,6-P₂ase, though predominantly cytosolic, were also present in the amyloplast. Based on distribution of enzymes, a probable pathway for starch biosynthesis in amyloplasts of developing wheat grains has been proposed.     

Carbonisation and morphological changes in modern dehusked and husked Triticum dicoccum and Triticum aestivum grains

Modern Triticum dicoccum and Triticum aestivum grains, with and without glumes, were subjected to experimental carbonisation under anoxic conditions. Experimental variables were the presence or absence of glumes, temperature, exposure time and heating rate. The maximum temperature was 600°C, the time of exposure was 60 min and the heating rate between 1 and 100°C/min. Length, width, area, mass loss and reflectance of uncarbonised and carbonised grains were measured as a function of the variables. The main effects of charring are an increase in width, decrease in length and formation of protrusions. Reflectance measurements allow for the determination of the temperature at which carbonisation occurred. The occurrence of protrusions on the pericarp, longitudinal imprints in the pericarp and concave flanks are observed and discussed. The calculated shape factor 100L/W is a useful tool for distinguishing between T. dicoccum and T. aestivum grains in samples that contain at least thirty specimens, but for single grains this method is problematic.     

Molecular characterization of the fate of transgenes in transformed wheat (Triticum aestivum L.)

Molecular analysis of the transgenes bar and gus was carried out over successive generations in six independent transgenic lines of wheat, until the plants attained homozygosity. Data on expression and integration of the transgenes is presented. Five of the lines were found to be stably transformed, duly transferring the transgenes to the next generation. The copy number of the transgenes varied from one to five in the different lines. One line was unstable, first losing expression of and then eliminating both the transgenes in R3 plants. Although the gus gene was detected in all the lines, GUS expression had been lost in R2 plants of all but one line. Rearrangement of transgene sequences was observed, but it had no effect on gene expression. All the stable lines were found to segregate for transgene activity in a Mendelian fashion.