Cloning and sequence analysis of genes involved in erythromycin biosynthesis in Saccharopolyspora erythraea: sequence similarities between EryG and a family of S-adenosylmethionine-dependent methyltransferases.

Summary Treatment of tomato seeds with ethyl methanesulphonate (EMS) followed by allyl alcohol selection of M₂ seeds has led to the identification of one plant (B15-1) heterozygous for an alcohol dehydrogenase (Adh) null mutation. Genetic analysis and expression studies indicated that the mutation corresponded to the structural gene of the Adh-1 locus on chromosome 4. Homozygous Adh-1 null mutants lacked ADH-1 activity in both pollen and seeds. Using an antiserum directed against ADH from Arabidopsis thaliana, which crossreacts with ADH-1 and ADH-2 proteins from tomato, no ADH-1 protein was detected in seeds of the null mutant. Northern blot analysis showed that Adh-1 mRNA was synthesized at wild-type levels in immature seeds of the null mutant, but dropped to 25% in mature seeds. Expression of the Adh-2 gene on chromosome 6 was unaffected. The potential use of the Adh-1 null mutant in selecting rare transposon insertion mutations in a cross with mutable Adh-1 ⁺ tomato lines is discussed.     

Linkage and cytogenetic maps of genes controlling endosperm storage proteins and isozymes in rye (Secale cereale L.)

Summary An F₁ plant fromSecale cereale ssp.ancestrale xtelocentric substitution lines3R of the cultivated rye Petkus spring was used as female in a cross with the inbred line Riodeva (I28), which has the standard chromosome arrangement. Single plants from this backcross progeny were analyzed for chromosome constitution, storage protein, and isozymic patterns. The seed protein loci were identified asSec-1a andSec-1b loci controlling 40-K-secalins and-secalins, respectively. These loci are located on the short arm of chromosome1R. TheSec-3 locus controlling high-molecular-weight secalins is located on the long arm of chromosome1R. A further seed protein locus,Pr-3 (55-K protein), was located on the short arm of chromosome1R. A linkage was found between the6Pgd-2 isozyme locus controlling 6-phosphogluconate dehydrogenase isozymes located on the long arm of chromosome1R and the four seed protein loci. The results favor the gene order:6Pgd-2 ...Sec-3 ... [centromere] ...Pr-3 ...Sec-1b ...Sec-1a. Other linkages detected werePer-3a andPer-3b (0.33±0.33 cM),Est-8 andEst-12 (0.33±0.33 cM), andGot-3 and centromere (20.57±2.42 cM). The proxidase (Per), glutamate oxaloacetate transaminase (Got), and esterase (Est) loci were located on chromosome arms2RS,3RL, and6RL, respectively. The distances and the maps obtained are compared with data available in the literature.     

Water-soluble proteins of mature barley endosperm: genetic control,polymorphism, and linkage with β-amylase and spring/winter habit

Summary Water-soluble proteins (WSP-2 and WSP-3) and -amylase (-AMY-1) were extracted from mature endosperms of 44 spring and 39 winter barley genotypes. The protein and enzyme isoforms were separated in isoelectric focusing gels with a pH gradient of 4–6.5. The Wsp-3 and -Amy-1 loci were located to chromosomes 4H using the wheat/barley chromosome addition lines. Segregation analysis of F₂ and doubled haploid populations showed Wsp-2 and -Amy-1 to be tightly linked, with a map distance of 11 cMorgans. Isoforms of WSP-2 possessed similar pIs to that of WSP-3 and overlapping bands were observed in the gels. These bands segregated independently in F₂ and doubled haploid populations, implying two unlinked genes. All three loci were found to be polymorphic: two alleles were detected at the Wsp-2 locus, three at Wsp-3 and two at -Amy-1. The frequency of alleles at all three loci was found to be different in winter and spring genotypes. Spring genotypes possessed a wider range of phenotypes than winter genotypes. Spring and winter genotypes could be distinguished on the basis of WSP-3 and - AMY-1 phenotypes. The linkage between Wsp-3 and -Amy-1 loci and genes controlling spring/winter habit on chromosome 4H is discussed. It is concluded that Wsp-3 and -Amy-1 can be used as genetic markers for spring/winter habit in barley genetic research and breeding.     

NADH dehydrogenase: a new molecular marker for homoeology group 4 in Triticeae. A map of the 4RS chromosome arm in rye

Summary Structural gene loci encoding the monomeric isozymes nicotin adenin dinucleotide dehydrogenase (NADH dehydrogenase or NDH) have been located on the 4AL, 4B, and 4DS chromosome arms of Triticum aestivum cv Chinese Spring, on the 4RS chromosome arm of Secale cereale cultivars Imperial, King II, Dakold, and Ailes, on the 4S¹ S/7S¹ chromosome of Aegilops longissima, the 4E of Elytrigia elongata, and the CSU-A of Aegilops umbellulata. All the results support the homoeologous relationships among these chromosomes in the five species studied. In addition, a map of the 4RS chromosome arm in cv Ailes has been realized, linking loci Pgm-1 (located on the 4RS chromosome arm) and Ndh-1 (17.91 cM), with an estimated distance between both loci and the centromere of 20.00 cM and 32.12 cM, respectively.     

Glutamine synthetase isoenzymes of Phaseolus vulgaris L.: subunit composition in developing root nodules and plumules

In the legume Phaseolus vulgaris L., glutamine synthetase (GS) (EC.6.3.1.2.) occurs as three cytosolic polypeptides, , and , and a plastidic polypeptide, . This paper describes the subunit composition of active octameric GS isoenzymes from root nodules and plumules using ionexchange high-performance liquid chromatography followed by two-dimensional denaturing gel electrophoresis and Western immunodetection. Root nodules contained four separable GS activities, three of which were composed mainly of cytosolic , / and GS polypeptides, whereas the fourth activity, consisted of plastidic GS polypeptides. The increase in GS activity during nodulation was due largely to the appearance of -containing isoenzymes, and to a lesser extent on the isoenzyme, whereas the -isoenzyme activity remained approximately constant throughout. Plumule GS from imbibed seeds was found to be composed of separate and isoenzymes, but 2 d after germination, plumule GS consisted of a mixture of , / and isoenzymes. The results from both nodules and plumules indicate that different cytosolic GS polypeptides in P. vulgaris are able to assemble into both homo-octameric and heterooctameric isoenzymes. Moreover, the changes in the patterns of isoenzymes observed during nodule development and plumule growth are interpreted to be caused both by temporal changes in the denovo synthesis of the polypeptides and also by their spatial separation in different cell types.Abbreviations 1D, 2D one-, two-dimensional - GS glutamine synthetase - GS_s GS semibiosynthetic activity - GS_t GS transferase activity - IEX-HPLC ion-exchange high-performance liquid chromatography - kDa kilodaltons - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis     

The chromosomal location of a third set of malate dehydrogenase loci,Mdh-3, in wheat,barley and related species

Summary A third set of malate dehydrogenase loci have been identified and located on the short arms of homoeologous group 5 chromosomes in wheat. Allelic differences have been found at each of the three Mdh-3 loci. However, Mdh-D3 appears to be least variable, with a second allele found only in Sears' Synthetic among a survey of 42 varieties. Homoeoloci were identified on chromosome 7 (5H) of Hordeum vulgare, the short arm of 5E in Agropyron elongatum and 5U in Aegilops umbellulata.     

The genetics of adult-plant blackleg (Leptosphaeria maculans) resistance from Brassica juncea in B. napus

The genetic control of adult-plant blackleg (Leptosphaeria maculans) resistance in a Brassica napus line (579NO48-109-DG-1589), designated R13 possessing Brassica juncea-like resistance (JR), was elucidated by the analysis of segregation ratios in F₂ and F₃ populations from a cross between R13 and the highly blackleg-susceptible B. napus cultivar Tower. The F₂ segregration ratios were bimodal, demonstrating that blackleg resistance in R13 was controlled by major genes. Analysis of the segregation ratios for 13 F₃ families indicated that blackleg resistance in these families was controlled by three nuclear genes, which exhibited a complex interaction. Randomly sampled plants of F₃ progeny all had the normal diploid somatic chromosome number for B. napus. The similarities between the action of the three genes found in this study with those controlling blackleg resistance in B. juncea is discussed.     

Block by 5-hydroxytryptamine of neuronal acetylcholine receptor channels expressed inXenopus oocytes

Summary 1. Effects of 5-hydroxytryptamine (5-HT) on neuronal nicotinic acetylcholine (ACh) receptor channels were investigated by expressing cloned channel subunits inXenopus oocytes.2. When channels were expressed with a combination of ₃ and ₄ subunits, 5-HT (10 to 300 µM) reversibly inhibited an inward current activated by 100 µM ACh in a concentration-dependent manner. The inhibition was also observed when ₃ subunit was combined with ₂ subunit instead of ₄ subunit, or ₄ subunit was combined with ₂ or _4–1 subunit instead of ₃ subunit to express channels.3. Compounds known to antagonize at 5-HT receptors (LY53857, metoclopramide and propranolol) exhibited an agonistic effect: they inhibited the ACh-activated current.4. The results suggest that 5-HT inhibits recombinant neuronal nicotinic receptor channels through a binding-site distinct from conventional 5-HT receptors. The binding-site may not be attributed to a unique type of channel subunits.     

Characterization of the MHC class II region in cattle. The number of DQ genes varies between haplotypes

The organization of the major histocompatibility complex (MHC) class II region in cattle was investigated by Southern blot analysis using human probes corresponding to DO, DP, DQ, and DR genes. Exon-specific probes were also employed to facilitate the assessment of the number of different bovine class II genes. The results indicated the presence of single DO and DR genes, at least three DR genes, while the number of DQ genes was found to vary between MHC haplotypes. Four DQ haplotypes, DQ ^{1 1} to DQ ^{2 4}, possessed a single DQ and a single DQ gene whereas both these genes were duplicated in eight other haplotypes, DQ ^{3 5} to DQ ^{9 12}. No firm evidence for the presence of bovine DP genes was obtained. The same human probes were also used to investigate the genetic polymorphism of bovine class II genes. DQ DQ , DR DR , and DO restriction fragment length polymorphisms (RFLPs) were resolved and in particular the DQ restriction fragment patterns were highly polymorphic. Comparison of the present result with the current knowledge of the class II region in other mammalian species suggested that the DO, DP, DQ, DR, and DZ subdivision of the class II region was established already in the ancestor of mammals. The DP genes appear to be the least conserved class II genes among mammalian species and may have been lost in cattle. The degree of polymorphism of different class II genes, as revealed by RFLP analyses, shows striking similarities between species.     

The Use of the [13C]/[12C] Ratio for the Assay of the Microbial Oxidation of Hydrocarbons

The study deals with a comparative analysis of the relative abundances of the carbon isotopes ¹²C and ¹³C in the metabolites and biomass of the Burkholderia sp. BS3702 and Pseudomonas putida BS202-p strains capable of utilizing aliphatic (n-hexadecane) and aromatic (naphthalene) hydrocarbons as sources of carbon and energy. The isotope compositions of the carbon dioxide, biomass, and exometabolites produced during the growth of Burkholderia sp. BS3702 on n-hexadecane (¹³C = –44.6 ± 0.2) were characterized by the values of ¹³C_{CO 2} = –50.2 ± 0.4, ¹³C_biom = –46.6 ± 0.4, and ¹³C_exo = –41.5 ± 0.4, respectively. The isotope compositions of the carbon dioxide, biomass, and exometabolites produced during the growth of the same bacterial strain on naphthalene (¹³C = –21 ± 0.4) were characterized by the isotope effects ¹³C_{CO 2} = –24.1 ± 0.4, ¹³C_biom = –19.2 ± 0.4, and ¹³C_exo = –19.1 ± 0.4, respectively. The possibility of using the isotope composition of metabolic carbon dioxide for the rapid monitoring of the microbial degradation of petroleum hydrocarbons in the environment is discussed.     

Monotelodisomics in pearl millet

Summary In the progeny of crosses between plants with the chromosome number 2n=13+2 telocentrics as the male parents and the normal diploids of Pennisetum typhoides S. & H., two plants with 2n=13+1 telocentric chromosome were located. These two plants were heterozygous for an interchange, since at diakinesis and metaphase I associations of four chromosomes were observed. These plants had a chromosome constitution of 2n=13+t (or 6+tI); one chromosome of a homologous pair was represented by a telocentric chromosome so was monosomic for one arm, that is, these plants were monotelodisomics (Kimber and Sears, 1968).     

Electrophoretic analysis of the high-molecular-weight glutenin subunits of Triticum monococcum,T. urartu,and the A genome of bread wheat (T. aestivum)

Summary The high molecular weight (HMW) subunit composition of glutenin was analysed by sodium dodecyl sulphate, polyacrylamide gel electrophoresis (SDS-PAGE) in the A genome of 497 diploid wheats and in 851 landraces of bread wheat. The material comprised 209 accessions of wild Triticum monococcum ssp. boeoticum from Greece, Turkey, Lebanon, Armenia, Iraq, and Iran; 132 accessions of the primitive domesticate T. monococcum ssp. monococcum from many different germplasm collections; one accession of free-threshing T. monococcum ssp. sinskajae; 155 accessions of wild T. urartu from Lebanon, Turkey, Armenia, Iraq, and Iran; and landraces of T. aestivum, mainly from the Mediterranean area and countries bordering on the Himalayan Mountains. Four novel HMW glutenin sub-units were discovered in the landraces of bread wheat, and the alleles that control them were designated Glu-Ald through Glu-Alg, respectively. The HMW subunits of T. monococcum ssp. boeoticum have a major, x subunit of slow mobility and several, less prominent, y subunits of greater mobility, all of which fall within the mobility range of HMW subunits reported for bread wheat. In T. monococcum ssp. monococcum the range of the banding patterns for HMW subunits was similar to that of ssp. boeoticum. However, two accessions, while containing y subunits were null for x subunits. The single accession of Triticum monococcum ssp. sinskajae had a banding pattern similar to that of most ssp. boeoticum and ssp. monococcum accessions. The HMW subunit banding patterns of T. urartu accessions were distinct from those of T. monococcum. All of them contained one major x and most contained one major y subunit. In the other accessions a y subunit was not expressed. The active genes for y subunits, if transferred to bread wheat, may be useful in improving bread-making quality.     

The addition of Dasypyrum villosum (L.) Candargy chromosomes to durum wheat (Triticum durum Desf.)

Summary Six monosomic addition lines were produced in which different Dasypyrum villosum (L.) Candargy chromosomes were added to the chromosome complement of Triticum durum Desf. cv. Creso. Each added alien chromosome was found to have a specific effect on plant morphology and fertility. Transmission rate varied widely (from 7.5 to 27.7%) among the six univalent chromosomes. Different monotelosomic addition plants derived by a relatively high frequency of chromosome misdivision were isolated. The addition lines should be useful for studying Dasypyrum chromosome homoeology and the introduction of alien variation into durum and common wheats.Research supported by a grant from the Italian Research Council for Finalized Project IPRA. Sub-project Plant Breeding, Paper No. 1095     

Ibf-1 (Iodine binding factor), a highly variable marker system in the Triticeae

Summary Isoelectric focusing of extracts from the endosperm of mature grains of hexaploid wheat and related species was used to study the genetic control of Iodine binding factor (IBF). Ten IBF bands were present in Chinese Spring (CS) and analysis of the nullisomictetrasomic and ditelosomic lines of CS showed nine of them to be controlled by genes on the long arms of the homoeologous group 5 chromosomes. Five alleles were detected at Ibf-A1 locus, four at Ibf-B1 and four at Ibf-D1 among a sample of 46 wheat genotypes. Homoeoloci were found on chromosome 5R of Secale cereale, 5E of Agropyron elongatum, 5U of Aegilops umbellulata, 5Agⁱ of Agropyron intermedium, 5S¹ and 4S¹ of Aegilops sharonensis and 4H of Hordeum vulgare.     

Genetic mapping of a quantitative trait locus (QTL) that enhances the shoot differentiation rate in Hordeum vulgare L.

A quantitative trait locus (QTL) controlling shoot differentiation from immature embryo callus was identified by linkage analysis with morphological and isozyme markers in barley, Hordeum vulgare L. Immature embryos were isolated from cvs Azumamugi (difficult to differentiate), Kanto Nakate Gold (easy to differentiate), their hybrids (F₁) and a backcross population derived from a cross Azumamugi x F₁. The embryos were cultured in vitro for callus initiation and subsequent shoot differentiation. The shoot differentiation rate was closely associated with ear type (v locus), isocitrate dehydrogenase isozyme (Idh-2), and esterase isozyme (Est-11). These markers were found to reside in a chromosome segment of approximately 30cM on chromosome 2. Recombination frequency was 9.9% between v and a proposed QTL named Shd1 (shoot differentiation), 11.5% between Idh-2 and Shd1, and 21.3% between Est-11 and Shd1. All data showed the Idh-2, v, Shdl and Est-11 loci to be arranged in this order from proximal to distal on the long arm of chromosome 2.     

Linkage relationships in the bovine MHC region. High recombination frequency between class II subregions

Class II genes of the bovine major histocompatibility complex (MHC) have been investigated by Southern blot analysis using human DNA probes. Previous studies revealed the presence of bovine DO , DQ , DQ , DR and DR genes, and restriction fragment length polymorphisms for each of these genes were documented. In the present study, the presence of three additional class II genes, designated DZ , DY , and DY , are reported. DZ was assumed to correspond to the human DZ gene while the other two were designated DY because their relationship to human class II genes could not be firmly established. The linkage relationships among bovine class II genes and two additional loci, TCP1B and C4, were investigated by family segregation analysis and analysis of linkage disequilibrium. The results clearly indicated that all these loci belong to the same linkage group. This linkage group is divided into two subregions separated by a fairly high recombination frequency. One region includes the C4, DQ , DQ , DR and DR loci and the other one is composed of the DO DY , DY , and TCPIB loci. No recombinant was observed within any of these subregions and there was a strong or fairly strong linkage disequilibrium between loci within groups. In contrast, as many as five recombinants among three different families were detected in the interval between these subregions giving a recombination frequency estimate of 0.17 ± 0.07. The fairly high recombination frequency observed between class 11 genes in cattle is strikingly different from the corresponding recombination estimates in man and mouse. The finding implies either a much larger molecular distance between some of the bovine class II genes or alternatively the presence of a recombinational hot spot in the bovine class II region.     

How polymorphic are class II loci of the mouse H-2 complex?

DNA was isolated from 75 mouse strains carrying classical H-2 haplotypes as well as haplotypes derived from wild mice. The DNA was digested with three restriction endonucleases, Bst EII, Eco RI, and Bam HI, the digests hybridized, using the Southern blotting technique, with probes for the class II genes A , A, E, and E , and the restriction fragment length polymorphism at these loci determined. The analysis revealed that the most polymorphic of the four loci is A , followed by E , and, at a different level, by E and A . There is a large difference in the degree of polymorphism between the A and E genes, on the one hand, and the A and E genes, on the other hand. There is no difference in the degree of polymorphism between the A and E genes. These findings do not substantiate previous postulates of a high A polymorphism and they do not agree with the hypothesis that the class II region is divided into highly polymorphic centromeric and less polymorphic telomeric subregions. Rather, it appears that the differences in the degree of polymorphism of the different segments of the class II region are determined by the class II loci themselves. The polymorphism of the less polymorphic class II genes is, however, still greater than the polymorphism of certain other genes on chromosome 17, notably the 4-globin pseudogene. The distribution of polymorphisms at the A and E loci suggests that even populations occupying relatively small geographical regions differ in alleles at these loci. Sharing of A alleles between unrelated populations is yet to be detected. A certain degree of linkage disequilibrium exists among the A , A , and E loci; by contrast, the E locus appears to vary largely independently of the other class II loci.     

Studies on aneuploids of Petunia

Summary The seven possible primary trisomics of Petunia (2 n= 14) located in the progenies of triploid, hypertriploid and hypotriploid plants were distinguished from one another and from diploid on the basis of cytological and morphological criteria. They were provisionally named as Oval, Semi, Slender, Pseudonormal, Arrow, Narrow and Giant. In three of the trisomics, the extra chromosome was identified for the first time at pachytene stage. Postpachytene studies revealed no precise relationship between the length of extra chromosome and the frequency of multiple association.     

Identification of RFLP markers linked to the cereal cyst nematode resistance gene (Cre) in wheat

The cereal cyst nematode (CCN) (Heterodera avenae Woll.) is an economically damaging pest of wheat in many of the worlds cereal growing areas. The development of CCN-resistant cultivars may be accelerated by the use of molecular markers. The Cre gene of the wheat line AUS 10894 confers resistance to CCN. Using a pair of near-isogenic lines (NILs) that should differ only in a small chromosome segment containing the Cre locus, we screened 58 group-2 probes and found two (Tag605 and CDO588) that detect polymorphism between the NILs. Nulli-tetrasomic and ditelosomic lines confirmed that the restriction fragment length polymorphism (RFLP) markers identified were derived from the long arm of wheat chromosome 2. Crosses between AUS 10894 and Spear and the NIL AP and its recurrent parent Prins were used to produce F₂ populations that gave the expected 31 segregation ratio for the resistance gene. Linkage analysis identified two RFLP markers flanking the resistance gene. Xglk605 and Xcdo588 mapped 7.3 cM (LOD=6.0) and 8.4 cM (LOD=6.7), respectively, from the Cre locus.