Durum Plants Research Articles

Introgression of the bread wheat D genome encoded Lr34/Yr18/Sr57/Pm38/Ltn1 adult plant resistance gene into Triticum turgidum (durum wheat)

Key MessageLack of function of a D-genome adult plant resistance gene upon introgression into durum wheat.The wheat Lr34/Yr18/Sr57/Pm38/Ltn1 adult plant resistance gene (Lr34), located on chromosome arm 7DS, provides broad spectrum, partial, adult plant resistance to leaf rust, stripe rust, stem rust and powdery mildew. It has been used extensively in hexaploid bread wheat (AABBDD) and conferred durable resistance for many decades. These same diseases also occur on cultivated tetraploid durum wheat and emmer wheat but transfer of D genome sequences to those subspecies is restricted due to very limited intergenomic recombination. Herein we have introgressed the Lr34 gene into chromosome 7A of durum wheat. Durum chromosome substitution line Langdon 7D(7A) was crossed to Cappelli ph1c, a mutant derivative of durum cultivar Cappelli homozygous for a deletion of the chromosome pairing locus Ph1. Screening of BC1F2 plants and their progeny by KASP and PCR markers, 90 K SNP genotyping and cytology identified 7A chromosomes containing small chromosome 7D fragments encoding Lr34. However, in contrast to previous transgenesis experiments in durum wheat, resistance to wheat stripe rust was not observed in either Cappelli/Langdon 7D(7A) or Bansi durum plants carrying this Lr34 encoding segment due to low levels of Lr34 gene expression. Key message

Evaluating the impact of Rht hypomorphic mutations in durum wheat

AbstractIncreasing the yield of wheat (Triticum spp.) requires identifying new allelic combinations by crossing or by creating useful variation in yield limiting genes. Wheat yield is impacted by many factors, including tiller number and seeds per tiller, both of which are impacted by the Reduced height (Rht) gene. Durum wheat [T. turgidum L. subsp. durum (Desf.) van Slageren] varieties are either standard height, wild type for Rht (Rht‐B1a), or are semidwarf and carry the Rht‐B1b allele. Rht‐B1b increases productive tillers but can result in plants too short for easy harvest in the northern United States and shorter coleoptiles that reduce dry soil germination. In this study, durum plants varying for Rht alleles created by ethyl methanesulfonate (EMS) mutagenesis were studied to determine the impact of each allele upon agronomic and seed traits. The projects’ goal is to increase durum wheat yield through the development of a plant with height intermediate between current full‐height and semidwarf varieties. Experiments included field trials, coleoptile length and gibberellic acid (GA) responsiveness assays, and an in vitro test to determine the impact of each Rht mutation upon binding to Gibberellin Insensitive Dwarf1 (GID1). It was found that the Rht‐B1b‐E529K allele conferred plant height and coleoptile length intermediate between Rht‐B1b and Rht‐B1a containing plants, while two Rht‐A1 alleles had lesser impacts with trends toward intermediate‐height plants. The results of this research demonstrate that hypomorphic Rht alleles that alter Rht binding to GID1 may prove useful in optimizing durum wheat height to increase yield across different growing conditions.

Reducing Nitrogen Dosage in Triticum durum Plants with Urea-Doped Nanofertilizers.

Nanotechnology is emerging as a very promising tool towards more efficient and sustainable practices in agriculture. In this work, we propose the use of non-toxic calcium phosphate nanoparticles doped with urea (U-ACP) for the fertilization of Triticum durum plants. U-ACP nanoparticles present very similar morphology, structure, and composition than the amorphous precursor of bone mineral, but contain a considerable amount of nitrogen as adsorbed urea (up to ca. 6 wt % urea). Tests on Triticum durum plants indicated that yields and quality of the crops treated with the nanoparticles at reduced nitrogen dosages (by 40%) were unaltered in comparison to positive control plants, which were given the minimum N dosages to obtain the highest values of yield and quality in fields. In addition, optical microscopy inspections showed that Alizarin Red S stained nanoparticles were able to penetrate through the epidermis of the roots or the stomata of the leaves. We observed that the uptake through the roots occurs much faster than through the leaves (1 h vs. 2 days, respectively). Our results highlight the potential of engineering nanoparticles to provide a considerable efficiency of nitrogen uptake by durum wheat and open the door to design more sustainable practices for the fertilization of wheat in fields.

Metabolic acclimation of tetraploid and hexaploid wheats by cold stress-induced carbohydrate accumulation

Metabolic acclimation of plants to cold stress may be of great importance for their growth, survival and crop productivity. The accumulation carbohydrates associated with cold tolerance (CT), transcript levels for genes encoding related enzymes along with damage indices were comparatively studied in three genotypes of bread and durum wheats differing in sensitivity. Two (Norstar, bread wheat and Gerdish, durum wheat) were tolerant and the other, SRN (durum wheat), was susceptible to cold stress. During cold stress (−5°C for 24h), the contents of electrolyte leakage index (ELI) in Norstar and then Gerdish plants were lower than that of SRN plants, particularly in cold acclimated (CA) plants (4°C for 14days), confirming lethal temperature 50 (LT50) under field conditions. Increased carbohydrate abundances in the cases of sucrose, glucose, fructose, hexose phosphates, fructan, raffinose, arabinose resulted in different intensities of oxidative stress in bread (Norstar) plants compared to durum plants (SRN and Gerdish) plants as well as in CA plants compared to non-acclimated (NA) ones under cold, indicating metabolic/regulatory capacity along with a decrease in ELI content and enhanced defense activities. A significant decrease in these carbohydrates, particularly sucrose, under cold in NA plants showed an elevated level of cell damage (confirmed by ELI) compared to CA plants. On the other hand, an increase in hexose phosphates, particularly in NA plants, indicated sucrose degradation along with greater production of glucose and fructose compared to CA plants. Under such conditions, a significant increase in transcript levels of sucrose synthase and acidic invertase confirmed these results. Under cold, the high ABA-containing genotypes like Norstar and then Gerdish, which were obvious in CA plants, partly induced relative acclimation of cells for acquisition of CT compared to SRN. These results reveal an important role of carbohydrate metabolism in creating CT in durum wheats (particularly in Gerdish) as well as bread wheat with possible responsive components in metabolic and transcript levels.

Preceding crop and weed management history affect denitrification and denitrifier community structure throughout the development of durum wheat

Aside from negative impacts on crop yield, the role of weeds in agricultural systems, particularly with respect to below ground function, is not well understood. In this study, we utilised a mesocosm approach with soils collected from a long-term rotation study to compare legacy effects of previous flax (Linum usitatissimum L.) or canola (Brassica napus L.) and different levels of weediness established over a decade by different herbicide use patterns. Despite few significant differences in initial soil chemical properties, the impact of level of weediness superseded that of the preceding crop and altered the growth of durum wheat (Triticum durum L.) during early development and the temporal dynamics of denitrification and the nirK denitrifier communities. The impact of the presence of durum plants in the mesocosm experiment was relatively small, however, the presence of durum plants did modify the legacy effects of preceding crop and weediness on the denitrifier community. Differences in denitrifier community structure were due to differential presence/absence and differential abundance of multiple T-RFs. To our knowledge, this is the first study to investigate the effects of long-term weed management strategies on denitrification. Our results indicate that weediness contributes to priming effects and unexplained variation associated with these soil processes and that weed management history should be taken into consideration when determining soil function and soil microbial community dynamics in agricultural systems.

Registration of wheat lines carrying the partial stripe rust resistance gene Yr36 without the Gpc-B1 high grain protein content allele.

While the high-temperature adult plant resistance gene Yr36 represents a promising source of quantitative and potentially race non-specific resistance to wheat stripe rust (causal organism Puccinia striiformis Westend. f. sp. tritici), its tight linkage (0.3 cM) with the high-grain protein content gene Gpc-B1 may hinder its introgression in certain cases, such as in soft wheat varieties requiring low grain protein content or in lines where the Gpc-B1 allele may be associated with a yield penalty. The development and registration of two donor lines, one tetraploid (Triticum turgidum L. ssp. durum; PI 656793) and one hexaploid (T. aestivum L. ssp. aestivum; PI 664549), each carrying the resistant wild emmer (T. turgidum ssp. dicoccoides) allele for Yr36 linked with the non-functional Gpc-B1 allele, are intended to overcome this potential limitation. Meiotic recombination events breaking the linkage between these two genes were discovered during the systematic screening of a population of 4,500 F2 durum plants (cv. Langdon background) used to fine map Yr36. One of the critical recombination events was selected for fixation by self-pollination and transferred to a California adapted spring hexaploid background (breeding line UC11105+10) through five generations of backcrossing. Genotypic and phenotypic data confirm the presence of Yr36 and the non-functional Gpc-B1 allele in both registered lines.

Involvement of root ABA and hydraulic conductivity in the control of water relations in wheat plants exposed to increased evaporative demand

We studied the possible involvement of ABA in the control of water relations under conditions of increased evaporative demand. Warming the air by 3°C increased stomatal conductance and raised transpiration rates of hydroponically grown Triticum durum plants while bringing about a temporary loss of relative water content (RWC) and immediate cessation of leaf extension. However, both RWC and extension growth recovered within 30min although transpiration remained high. The restoration of leaf hydration and growth were enabled by increased root hydraulic conductivity after increasing the air temperature. The use of mercuric chloride (an inhibitor of water channels) to interfere with the rise on root hydraulic conductivity hindered the restoration of extension growth. Air warming increased ABA content in roots and decreased it in shoots. We propose this redistribution of ABA in favour of the roots which increased the root hydraulic conductivity sufficiently to permit rapid recovery of shoot hydration and leaf elongation rates without the involvement of stomatal closure. This proposal is based on known ability of ABA to increase hydraulic conductivity confirmed in these experiments by measuring the effect of exogenous ABA on osmotically driven flow of xylem sap from the roots. Accumulation of root ABA was mainly the outcome of increased export from the shoots. When phloem transport in air-warmed plants was inhibited by cooling the shoot base this prevented ABA enrichment of the roots and favoured an accumulation of ABA in the shoot. As a consequence, stomata closed.

Spontaneous haploids in durum wheat: their cytogenetic characterization

Durum or macaroni wheat (Triticum turgidum L., 2n = 4x = 28; AABB genomes) arose as a natural hybrid between two wild species, Aegilops speltoides Tausch (2n = 2x = 14; BB genome) and Triticum urartu Tumanian (2n = 2x = 14; AA genome). The two progenitors hybridized in nature about half a million years ago and gave rise to tetraploid wheat, presumably in one step as a result of functioning of unreduced gametes in their hybrid BA (amphihaploid). It is easily possible to go back on the evolutionary scale and obtain durum haploids BA, and then regenerate tetraploid durum plants from them. Interestingly, such a reversion to haploidy does occur in nature as well, although at a very low frequency. This article reports on the occurrence of two spontaneous durum haploids and describes their chromosomal characteristics. The haploids (euhaploids, to be precise) had 14 somatic chromosomes, which, on fluorescent genomic in situ hybridization (fl-GISH), could be distinguished as 7 A-genome and 7 B-genome chromosomes. At meiosis, only 2.3 and 2.7% of the chromosomes paired in the two haploids, because of the presence of the homoeologous pairing-suppressor gene, Ph1. The Ph1-induced lack of pairing is a prerequisite for chromosome doubling through the formation of unreduced gametes that give rise to tetraploid durum wheats.

Identification and mass spectrometric characterization of glycosylated flavonoids in Triticum durum plants by high‐performance liquid chromatography with tandem mass spectrometry

A mass spectrometric method for extensive detection and semi-quantitative determination of flavonoid glycosides in stem and leaves of young Triticum durum plants is presented. About 100 g of sample were lyophilized and ground, and the compounds of interest were then extracted, cleaned-up, and fractionated using high-performance liquid chromatography (HPLC). Tandem mass spectrometry analyses were performed using a quadrupole-linear ion trap instrument with an information-dependent data acquisition (IDA) protocol that looped two experiments, enhanced MS scan and enhanced product ion scan. Various glycoconjugates, which are all derivatives of only four flavones, apigenin, luteolin, chrysoeriol and tricin, were identified and belong to the following categories: 7 monoglycosides, 31 diglycosides, 15 triglycosides and 1 tetraglycoside. Among these some acylated glycosides were found. Tricin derivatives are present exclusively as O-glycosides, while apigenin and luteolin are present always as C-glycosides. Semi-quantitative estimation was performed by using the monoglycoside and diglycoside of quercetin as internal standards.

Nitrogen and Carbon Isotope Composition of Wheat Grain: Alteration due to Sink-source Modifications at Flowering

An experiment of sink-source modifications was performed in field-grown wheat at flowering on five varieties (four Triticum aestivum and one T. durum); among these, two varieties were non-awned. The sink-source modifications were blade removal (treatment B) and blade deletion and stalk and sheaths darkening (treatment C). Intact plants (treatment A) acted as a control. In the three awned varieties, the effect of awn removal was also analysed. Two lots of tests were initially performed, with awns (treatments A+, B+, C+) or with awns removed (treatments A−, B−, C−). Yields and carbon and nitrogen isotope compositions were then determined on seeds harvested at maturity. Blade deletion reduced the yield by one third whereas additional stalk and sheaths darkening led to a two thirds decrease. Nitrogen yield was similarly affected but the deprivation due to treatment C was less marked for nitrogen than for carbon. Thus, treatment C produced grains which were notably richer in nitrogen than plants which had received treatment A and B. For each of the five varieties, the decrease in yield was highly correlated with a 13C enrichment of grains (roughly from −28 to −25‰). In contrast with the 13C variations, there was no general trend for 15N evolution among treated plants. The final yield for treated T. durum plants was strongly modified when the awns were removed at flowering whereas no significant variations were noted in the other T. aestivum awned varieties. T. durum seeds obtained from plants deprived of awns were notably depleted in 13C and in 15N compared to control plants. Isotope compositions of C and N of the different pools supplying the grain filling were estimated and discussed in the context of the physiology of the mother plants between flowering and maturity.

A gene for embryo–endosperm compatibility and seed viability in alloplasmic Triticum turgidum

Alien cytoplasms have more deleterious effect on Triticum turgidum L. var. durum (2n = 28; 14 II; AABB) than on T. aestivum L. (2n = 42; 21 II; AABBDD). The species cytoplasm specific (scs) nuclear genes from the cytoplasm donor species and scs homoeoalleles ameliorate cytoplasmic effects in alloplasmic wheats. The D-genome chromosome(s) of T. aestivum must have a scs gene(s) that is absent in T. turgidum. Also, the dosage of scs homoeoalleles must be more favorable for nucleocytoplasmic compatibility (NCC) in 6x than 4x wheats. This paper reports that two genes, scs on 1DL telosome from T. aestivum 'Selkirk' and a vitality (Vi) gene from (Aegilops cylindrica) T. aestivum 'Selkirk', restored nucleocytoplasmic and embryo–endosperm compatibility, fertility, seed viability, and plant vigor in Ae. squarrosa and Ae. cylindrica 29-chromosome durum plants. In the absence of Vi, 29-chromosome plants set a few sporadic plump seed in selfed spikes but a few plump (about same number as in selfed spikes) and a large number of shrivelled seed in spikes crossed with euplasmic durum. Seed with Vi embryos were viable and those without Vi were nonviable. Similarly, Vi restored fertility to spontaneous aneuploids in the progeny of Ae. squarrosa 29-chromosome plants. Euploid sibs with Vi, but without 1DL telosome, had greatly reduced vigor. The control 'Selkirk' and durum wheats did not have Vi. Hence, Vi may have originated by mutation in Ae. cylindrica 'Selkirk' or Ae. cylindrica durum. Similar spontaneous changes may improve fertility in interspecific hybrid progeny and condition complex inheritance of fertility restoration in alloplasmic wheats.Key words: T. aestivum, Vi and scs genes, xenia effect, zygotic sterility, seed viability, nucleocytoplasmic compatibility, syndrome of cytoplasmic effects.

Transfer of a species cytoplasm specific (scs) gene from Triticum timopheevii to T. turgidum

Initial attempts to substitute euploid nuclei of Triticum turgidum L. or T. aestivum L. into Aegilops longissima S. &L. cytoplasm failed because an alien chromosome remained fixed in the Triticum nucleus. The alien chromosome had gene(s) conditioning sporophytic sterility (also known as the gameticidal or Cuckoo effect). Subsequently, an exceptional 29-chromosome, male-sterile plant with spontaneously improved female fertility was used as a source of Ae. longissima cytoplasm, and a fully fertile alloplasmic common wheat 'Selkirk' line was developed. However, alloplasmic 'Selkirk' crossed with durum wheat as a recurrent male parent did not produce euploid plants. Instead, chromosome 1D or telocentric 1DL of 'Selkirk' was retained and male-sterile plants with 29 chromosomes were obtained. They set two seed types: a few that were plump and viable (PVi) and a large number that were shrivelled and inviable (SIv). The 1DL was deleted by crossing these plants to T. timopheevii, backcrossing the F1's to T. timopheevii, and repeatedly backcrossing the timopheevii-like plants to durum as the recurrent male parent. The resulting euploid durum plants with Ae. longissima cytoplasm were male sterile and set a 1:1 ratio of PVi and SIv seeds. Thus, a species cytoplasm specific (scs) gene of T. timopheevii was transferred to durum and caused male sterility and abortion of embryos lacking this gene. In conclusion, (i) the scs gene was expressed as a dominant sterility gene, restored seed viability, and partial compatibility between the durum nucleus and Ae. longissima cytoplasm and (ii) a scs gene on 1DL also caused dominant sterility in durum but not in alloplasmic common wheat. Hence, alien scs homoeoallele(s) conditioned sterility and seed abortion in alloplasmic durum but not in T. aestivum and T. timopheevii.Key words: interspecific nucleocytoplasmic genetics, sporophytically controlled sterility, B-genome donor, scs gene.

The presence of three groups of Scalavatis and other hexaploid bread wheat plants contaminating durum wheat fields in Cyprus

The mutual resemblance of 81 hexaploid wheat plants contaminating durum fields on Cyprus was investigated by determining their phenotypes of gliadins, glutenins and peroxidase. Three groups were generated by clustering programs. Group 1 consists of plants with ears resembling those of durum plants, plants of groups 2 and 3 have a bread wheat appearance. Suggestions on the origin of these three groups are made. Group 1 may derive from (back)crosses between hexaploid and durum plants since time immemorial. If so, the hexaploid plants would have served as ‘donor’ of the D genomes, and durum plants as the ‘recurrent parent’. Groups 2 and 3 may derive from two introductions, either from two countries, or from the same country, but then in different periods.

Plant part selection and evaluation of factors affecting analysis and recovery of nitrate in irrigated durum wheat tissue

Abstract Periodic nitrate tissue tests are used to determine nitrogen (N) fertility status of a variety of crops. Data on the importance of plant part selection, sample handling techniques, grinding criteria and extraction conditions in N03‐N analysis of wheat tissue are essential if the procedure is to achieve widespread adoption and use. Detailed partitioning of field grown durum spring wheat (Triticum durum) plants at the Feekes 2 (3–4 leaf), 6 (joint) and 10 (boot) growth stages was conducted to document which plant part exhibits the greatest accumulation of NO3‐N. Sample handling, fineness of tissue grinding, different tissue: extractant ratios and extraction times were examined to determine their effects on NO3‐N recovery. Partitioning data confirmed previous findings which identified the basal stem between ground level and the seed prior to jointing and the 5 cm of stem just above ground level thereafter as the plant parts showing the greatest accumulation of NO3‐N. Therefore, these plant parts hav...

The chromosomal location of a gene (msg) affecting megasporogenesis in durum wheat

An aneuploid durum wheat line (Triticum turgidum L. var. durum) having 13 chromosome pairs and 2 unpaired chromosomes at metaphase I of meiosis in pollen mother cells (i.e., monosomic for chromosomes 7A and 7D) was observed to produce some progeny plants with 2n = 40 chromosomes. These aneuploid (triploid) plants were usually weak and sterile. Triploid plants also occurred in the progeny of durum plants monosomic for chromosome 7A, or in progeny of plants that were mono-telodisomic or ditelomonotelosomic for chromosome 7Aq (13 II + 1 t II or 13 II + t II + t I) but not in the progeny of plants ditelomonotelosomic for chromosome 7Ap (13 II + t II + t 1). Therefore, there is a gene(s) on chromosome arm 7Ap that prevents the production of diploid (2n) egg cells in wheat. In the absence of 7Ap, a portion of the egg cells have 26 chromosomes, which when fertilized with a pollen nucleus with 14 chromosomes, produces progeny plants with 2n = 40 chromosomes. The data also indicated that chromosome arm 7Dp probably contains a second gene that is capable of preventing the production of triploid plants. Key words: Triticum turgidum L. var. durum, polyploidy, aneuploid, triploid, monosomic.

CYTOPLASMIC HOMOLOGY BETWEEN AEGILOPS TRIUNCIALIS AND AE. UMBELLULATA

The Triticum aestivum L. (2n = 42; AABBDD) and T. durum Desf. (2n = 28; AABB) genomes were substituted into the cytoplasm of two Aegilops triuncialis L. (2n = 28; CCCuCu) accessions. In F1 (ABCCu) and in a 42-chromosome backcross plant (AABBCCu) meiotic pairing between the C- and Cu-genome chromosomes was substantially reduced. Therefore, some of the pairing in the F1 of Ae. caudata L. (2n = 14; CC) × Ae. umbellulata Zhuk. (2n = 14; CuCu) may have been of the homoelogous type. In early backcrosses with T. aestivum, the euploid gametes or zygotes did not function in plants having Aegilops chromosome(s), but euploid plants with Ae. triuncialis cytoplasm were obtained in the fifth backcross. These plants were used as the cytoplasmic source to substitute the T. durum genomes into Ae. triuncialis cytoplasm; euploid durum plants were obtained in the second backcross. Alloplasmic T. durum and T. aestivum plants with Ae. triuncialis cytoplasm had closer resemblance to the plants with Ae. umbellulata cytoplasm than to those with Ae. caudata cytoplasm. These nucleo-cytoplasmic interactions indicated cytoplasmic similarity between Ae. umbellulata and Ae. triuncialis.

MALE STERILITY-FERTILITY RESTORATION SYSTEMS IN TRITICUM DURUM

Triticum durum Desf. selection 56-1 (2n = 28,AABB) genomes were introduced into the cytoplasms of six species of Triticum, 14 of Aegilops, and one each of Secale and Haynaldia by the backcross method. Of the 22 alloplasmic lines, 14 were completely male sterile, four were partially fertile, and the remaining four, having cytoplasms of T. dicoccoides Körn, Ae. kotschyi Boiss., Ae. variabilis, Eig. or H. villosa L., were of normal fertility. Eleven of these lines headed late and 14 had lower plant height than the control euplasmic T. durum. The 14 male-sterile lines were crossed with five R-lines derived from crosses involving T. nudiglumis Nabalek and T. durum, and male-fertility restoration in F1 hybrids was examined. All five R-lines restored male fertility to six of the male-sterile lines having cytoplasms of Ae. speltoides Tausch., Ae. bicornis Forsk., T. nudiglumis, T. araraticum Jakubz., T. timopheevi Zhuk., or T. zhukovskyi Men. &Er. The male fertility of the alloplasmic line having rye (secale cereale L.) cytoplasm was completely restored by RE 5 and partially by RE 2. Also, RE 5 restored plant vigor to durum plants having rye cytoplasm. None of the five R-lines restored male fertility or plant vigor to any of the six male-sterile lines having cytoplasms of T. boeoticum Boiss., Ae. caudata L., Ae. umbellulata Zhuk., Ae. heldreichii Holzm., Ae. sharonensis Eig., or Ae. triaristata Willd. In general, F2 data from crosses involving four of the male-sterile lines and five R-lines indicated that male fertility restoration was simply inherited.

Survival and chlorophyll mutation in Triticum durum plants raised from aged seeds

Survival and chlorophyll mutation in Triticum durum plants raised from aged seeds

Studies on Restoration and Substitution of Nucleus (Genome) in <i>Aegilotricum</i>

1) Abnormal stomata, which appeared in the variegated leaves of cytoplasmic male-sterile durum plants were investigated cytologically and classified in six types according to two criteria, namely; degree of differentiation of guard cells and their nuclei, and abnormality or deficiency of subsidiary cells.2) Observation on the cross-section of variegated leaves revealed that the inner air space of the abnormal stomata was smaller than that of normal stomata and that palisade cells in the variegated portion were rather round and arranged irregularly.3) The frequency of occurrence of abnormal stomata was investigated. The percentage of abnormal stomata in the variegated portion reached as high as 38%, but the mean value in the variegated leaf was 12% in longitudinal and 9% in transverse directions. Mature leaves of male-sterile plants do not have abnormal stomata any longer (only 0.2% of the stomata was abnormal).4) Alteration of stomatal distribution was observed in the variegated leaves, that is; the average number of stomata per unit length is decreased in longitudinal direction and is increased in transverse direction, as compared to the number found in normal green leaves.5) It was ascertained from the x2-test that abnormal stomata did not occur at random in the variegated leaf, but occurred serially to a certain degree and that more series were found in longitudinal than in transverse direction.6) Cell length and nuclear length of guard cells in the variegated leaf varied widely and decreased in comparison with those of the normal green leaf. On the other hand, some subsidiary cells were increased in length (although the mean value of cell length was decreased). Width of cell and nucleus did not change to the same degree.7) A correlation between cell length and nuclear length of guard cells was found expressed by a correlation coefficient as high as +0.9137.8) The data presented lead to the assumption that an unknown cytoplasmic factor induces abnormal metabolism in winter leaves, leading to abnormal development of stomata, and on the other hand, to formation of abnormal chloroplasts. The abnormal stomata seem to accelerate the chlorophyll variegation.The present author wishes to express his cordial thanks to Dr. H. G. duBuy, National Institute of Allergy and Infectious Diseases, U.S.A., who was kind enough to read over the manuscript and make some corrections in the style.

Studies on Restoration and Substitution of Nucleus in <i>Aegilotricum</i>

Exchange of nucleus by means of successive backcrosses to both parents has been carried out in an experiment with Tschermak's amphidiploid hybrid _??_ Aegilops ovata×Triticubr durum, known under the name of Aegilotricum.In the restoration backcrosses to Ae. ovata the process of restoring the ovata genomes to the ovata plasma has been accomplished in the course of three successive crosses. The final offspring were morphologically similar to Ae. ovata, had 14 bivalents at first metaphase and normal anthers filled with functional, good pollen.In the substitution backcrosses of Aegilotricum to T. durum, the three successive crosses resulted in durum plants with ovata plasma. They had at first metaphase 14 completely paired bivalents; the tetrad formation was normal but later all pollen was degenerating, and the small dry anthers contained empty pollen. However, the female organs were fully functional and seed setting by artificial pollination with durum or dicoccum pollen was normal. The male sterility was transmitted to the next generation by the mother.Similar pollen degeneration was observed in normal T. durum which have matured in winter after vernalization and long-day treatments.It is further to be noticed that durum plants with ovata plasma having extra chromosomes of Ae. ovata as univalents in addition to the 14 duram bivalents were only partially male-sterile. Especially, a 29-chromosome plant showed a very high pollen fertility of 92%.