The sequential combination of C-banding and in situ hybridization techniques applied in this or i... more The sequential combination of C-banding and in situ hybridization techniques applied in this or in a reverse order, are used to recognize targeted chromosomal regions in cereals. Both methods are described whereby standard chromosome squash preparations are followed by: i) C-banding technique using Leishman stain and a slightly modified in situ hybridization technique using biotin-labeled DNA probes, or ii) fluorescence in situ hybridization technique and C-banding. Both approaches have been successfully used onto mitotic chromosomes of rye and wheat resulting suitable for both their identification and detection of targeted sites.
Hordeum brachyantherum Nevski includes two subspecies: the diploid (2×) subsp. californicum, and ... more Hordeum brachyantherum Nevski includes two subspecies: the diploid (2×) subsp. californicum, and subsp. brachyantherum, which itself includes a tetraploid (4×) and a hexaploid (6×) cytotype. The phylogenetic relationships between these taxa and the origin of the polyploids remain controversial. To provide additional information to the many molecular phylogenetic analyses conducted within Hordeum, FISH-based karyotypes were produced for all subspecies/cytotypes within H. brachyantherum. Chromosomes of H. roshevitzii and H. marinum subsp. gussoneanum were also analysed since these species are potentially involved in the origin of the polyploids. For karyotyping, ten repetitive DNA sequences were screened to indentify repeats showing sufficient diversity in terms of copy number and localisation that they might serve as physical markers for distinguishing between each mitotic chromosome pair in all accessions. Genomic in situ hybridisation (GISH) was used to distinguish between subgenomes in polyploids. The karyotype maps allowed the assessment of the chromosomal diversity within species/cytotypes and the identification of possibly homoeologous chromosomes. The results show a wide divergence between the chromosomes of subsp. californicum and H. roshevitzii, and with their supposed derivatives in subsp. brachyantherum 4×. One of the three subgenomes of subsp. brachyantherum 6× is derived from subsp. gussoneanum with no genomic reorganisation (i.e., neither amplification nor loss of the repetitive DNA sequences analysed). It is generally accepted that subsp. brachyantherum 4× is the other progenitor of subsp. brachyantherum 6×, but the present results suggest this to be unlikely. The present findings thus show the cytogenetic diversity and genomic structure of H. brachyantherum, and reveal its complex evolutionary history, in which chromosomal diversification and allopolyploidy have played important roles.
... Likewise, H. chilense has been crossed with Secale species, obtaining hybrids with S. cereale... more ... Likewise, H. chilense has been crossed with Secale species, obtaining hybrids with S. cereale (FINCH & BENNET 1980, MARTIN & CUBERO 1981), S. africanum, S. montanum and S. vavilovii (FINCH & BENNETT 1980), and with 6x-Triticale (SCHRADER & POHLER 1985). ...
1991. Changes in expression of seed storage protein genes effected by chromosome 1D of wheat. Gen... more 1991. Changes in expression of seed storage protein genes effected by chromosome 1D of wheat. Genome, 34: 845-848. The effect of chromosome 1D of wheat on expression of seed storage protein genes was analyzed using a lR(1D) substitution line and derivatives of this line that had lost 1R (ID nullisomic). The results of one-and two-dimensional (two pH) electrophoresis suggested that nucleotide sequences located on ID were involved in induction and suppression of these protein genes and in posttranscriptional modification of polypeptides produced by the seed storage protein genes. The action attributed to ID appeared to affect only genes located on chromosome 6A.
Three repeated DNA sequences (rDNA 5S, 18S-5.8S-26S and telomeric repeats) were localised in the ... more Three repeated DNA sequences (rDNA 5S, 18S-5.8S-26S and telomeric repeats) were localised in the genomes ofLobelia brasiliensis andL. imperialis var.kanitzii (subg.Tupa), both with 2n = 28, by fluorescence in situ hybridization (FISH). The results were used to analyse the genomic relationship between the species. With probe pTa71, the karyotypes of these species showed only one NOR site. Probe pTa794, which
Simple Sequence Repeats (SSRs) are known to be scattered and present in high number in eukaryotic... more Simple Sequence Repeats (SSRs) are known to be scattered and present in high number in eukaryotic genomes. We demonstrate that dye-labeled oligodeoxyribonucleotides with repeated mono-, di-, tri, or tetranucleotide motifs (15-20 nucleotides in length) have an unexpected ability to recognize SSR target sequences in non-denatured chromosomes. The results show that all these probes are able to invade chromosomes, independent of the size of the repeat motif, their nucleotide sequence, or their ability to form alternative B-DNA structures such as triplex DNA. This novel and remarkable property of binding SSR oligonucleotides to duplex DNA targets permitted the development of a non-denaturing fluorescence in situ hybridization method that quickly and efficiently detects SSR-enriched chromosome regions in mitotic, meiotic, and polytene chromosome spreads of different model organisms. These results have implications for genome analysis and for investigating the roles of SSRs in chromosome structure and function.
The chromosome constitution and meiotic behaviour of 15 plants of F2 and BCI progenies from the h... more The chromosome constitution and meiotic behaviour of 15 plants of F2 and BCI progenies from the hybrid of 6xtriticale x T. turgidum (L.) ssp. turgidum cony. durum (Desf.) (Fl: 2n = 5x = 35; AABBR) were studied using Cbanding patterns of both wheat and rye chromosomes as identifying features. Patterns in PMCs at first metaphase permitted the individual identification of the chromosomes of nine of the wheat and all of rye in all plants. The external length and internal heterochromatin of chromosomes were analysed and found to influence the meiotic behaviour of each chromosome pair. The existence of effects of both arm length and amount and distribution of heterochromatin on pairing are assumed. The results seem to demonstrate that of the two mentioned factors the first is the most important.
Z-DNA forms transiently behind the active RNA polymerases, because of the mechanical torsional st... more Z-DNA forms transiently behind the active RNA polymerases, because of the mechanical torsional stress produced during transcription. In this paper, we explore the possibility that the distribution of Z-DNA stretches signals the sites related to nuclear transcription. To localize transcription, the in situ assay for active RNA polymerases, that allows the elongation of the already initiated transcripts but no initiation of new ones (run-on experiments), was carried out in isolated nuclei of Allium cepa L. root meristems. Both nucleolar and non-nucleolar sites appeared labelled. Nucleoli were most active in transcription than the multiple non-nucleolar foci altogether. In situ immunodetection of Z-DNA provided images that were comparable to those obtained after the run-on assay, with one exception: while Z-DNA and transcription sites were scattered throughout the whole nucleus, Z-DNA also accumulated in the nuclear periphery, where no transcription foci were detected in run-on assays. The peripheral Z-conformation signals might correspond to dsRNA segments present in the pre-mRNA in the process of their export to cytoplasm. The Z-containing structures nearly disappeared when non-nucleolar RNA polymerase II-dependent transcription had been previously abolished by the adenosine analogue DRB (5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole). This inhibition selectively decreased the amount of all nucleoplasmic Z structures. On the other hand, the inhibition of the nucleolar RNA polymerase I by cordycepin (3'-deoxyadenosine) prevented the presence of Z-DNA in nucleoli. We propose to use the in situ immunodetection of Z-DNA as a marker of the transcription level in both nucleolus and non-peripheral nucleoplasmic regions of nuclei. Co-detection of Z-DNA and of intermediate filament (IF) proteins, the major components of the nuclear matrix, was also carried out. The IFA antibody recognizes a conserved epitope essential for dimerization of the multiple IF proteins. They co-localized with most nucleolar Z-DNA, but not with the nucleoplasmic ones. In the nuclear periphery, the Z-positive signals were adjacent to the IF proteins constituting the lamina, though both signals did not often co-localize.
ABSTRACT Two intervarietal hybrids of common wheat, Triticum aestivum L., are meiotically analyze... more ABSTRACT Two intervarietal hybrids of common wheat, Triticum aestivum L., are meiotically analyzed using the C-banding staining method. The C-banding pattern of nine meiotic chromosomes (4A, 7A, and the seven of the B genome) permitted their unequivocal recognition at first metaphase plates. The pairing frequency of each B-genome chromosome arm was scored. Data on the pairing frequency of the arms, separately considered, are applied to calculate expected pairing of whole chromosomes and whole genomes. The application of mathematical models to predict the genome pairing using either equal or different frequencies per chromosome arm is discussed.Key words: meiotic analysis, Triticum aestivum L., C-banding.
SUMMARY - It is analyzed the primary structure of some of the high molecular weight (HMW) gluteni... more SUMMARY - It is analyzed the primary structure of some of the high molecular weight (HMW) glutenin genes and their degree of conservation in the genera Triticum, Secale and Aegilops. The primary structure presented information on the allelic composition and the quantities of the endosperm protein fractions (HMW, LMW and gliadins) in the doughs of different common wheat cultivars and lines in process of selection. Dough W is correlated with the quantity of HMW subunits present in the gluten and in particular with the quantity of type x HMW subunits. The gliadins and the LMW subunits appear to act as a "solvent" which modifies the rheological properties of the dough by interfering with the polymerization of the HMW subunits, or by altering the relative amounts of the different types of subunits available. The contribution of each type of subunit in the organization of the intermolecular links, the formation of the multiproteic aggregates, and the properties of the gluten, is...
El aborto es un tema muy espinoso y de complejo abordaje en el que fácilmente se pueden mezclar a... more El aborto es un tema muy espinoso y de complejo abordaje en el que fácilmente se pueden mezclar argumentos científicos o clínicos con otros de orden político o ideológico. De ahí la necesidad de partir de un razonamiento riguroso, honesto, y libre de todo tipo de prejuicio, error o engaño. Partiendo de esa premisa este trabajo se ha dividido en cuatro partes principales: una introducción en la que se hace una distinción entre un pensamiento riguroso y uno manipulador y las consecuencias que de ellos se derivan; una segunda parte en la que se hace una descripción del marco jurídico que definió las políticas de salud hoy vigentes en la materia; una tercera parte en la que se mide la evolución del aborto a lo largo de los últimos doce años, y finalmente, una última parte en la que se hace un análisis bioético del problema social que supone el aborto y de las consecuencias que se derivan de su práctica a nivel nacional.
TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik, 1984
The meiotic identification of nine pairs of chromosomes at metaphase I of meiosis of Triticum aes... more The meiotic identification of nine pairs of chromosomes at metaphase I of meiosis of Triticum aestivum (B genome, 4A and 7A) has been achieved using a Giemsa C-banding technique. As a result, the analysis of the pairing of each chromosome arm in disomic and monosomic intervarietal hybrids between 'Chinese Spring' and the Spanish cultivar 'Pané 247' could be carried out. Differences in the chiasmata frequencies per chromosome arm cannot be explained on the basis of relative arm lengths only. Possible effects of arm-to-arm heterochromatic differences on meiotic pairing are discussed.
SUMMARY - Most cereals have genomes with redundant genes arising from genome doubling or polyploi... more SUMMARY - Most cereals have genomes with redundant genes arising from genome doubling or polyploidy. Differences in the expression levels of duplicate genes have long been a subject of great interest since new genes can arise from redundant duplicates. Such analysis requires a technique that can discriminate between the expressions of two (or more) almost identical genes (homoeologues). In previous
Hordeum marinum is a species complex that includes the diploid subspecies marinum and both diploi... more Hordeum marinum is a species complex that includes the diploid subspecies marinum and both diploid and tetraploid forms of gussoneanum. Their relationships, the rank of the taxa and the origin of the polyploid forms remain points of debate. The present work reports a comparative karyotype analysis of six H. marinum accessions representing all taxa and cytotypes. Karyotypes were determined by analysing the chromosomal distribution of several tandemly repeated sequences, including the Triticeae cloned probes pTa71, pTa794, pAs1 and pSc119·2 and the simple sequence repeats (SSRs) (AG)10, (AAC)5, (AAG)5, (ACT)5 and (ATC)5. The identification of each chromosome pair in all subspecies and cytotypes is reported for the first time. Homologous relationships are also established. Wide karyotypic differences were detected within marinum accessions. Specific chromosomal markers characterized and differentiated the genomes of marinum and diploid gussoneanum. Two subgenomes were detected in the t...
The sequential combination of C-banding and in situ hybridization techniques applied in this or i... more The sequential combination of C-banding and in situ hybridization techniques applied in this or in a reverse order, are used to recognize targeted chromosomal regions in cereals. Both methods are described whereby standard chromosome squash preparations are followed by: i) C-banding technique using Leishman stain and a slightly modified in situ hybridization technique using biotin-labeled DNA probes, or ii) fluorescence in situ hybridization technique and C-banding. Both approaches have been successfully used onto mitotic chromosomes of rye and wheat resulting suitable for both their identification and detection of targeted sites.
Hordeum brachyantherum Nevski includes two subspecies: the diploid (2×) subsp. californicum, and ... more Hordeum brachyantherum Nevski includes two subspecies: the diploid (2×) subsp. californicum, and subsp. brachyantherum, which itself includes a tetraploid (4×) and a hexaploid (6×) cytotype. The phylogenetic relationships between these taxa and the origin of the polyploids remain controversial. To provide additional information to the many molecular phylogenetic analyses conducted within Hordeum, FISH-based karyotypes were produced for all subspecies/cytotypes within H. brachyantherum. Chromosomes of H. roshevitzii and H. marinum subsp. gussoneanum were also analysed since these species are potentially involved in the origin of the polyploids. For karyotyping, ten repetitive DNA sequences were screened to indentify repeats showing sufficient diversity in terms of copy number and localisation that they might serve as physical markers for distinguishing between each mitotic chromosome pair in all accessions. Genomic in situ hybridisation (GISH) was used to distinguish between subgenomes in polyploids. The karyotype maps allowed the assessment of the chromosomal diversity within species/cytotypes and the identification of possibly homoeologous chromosomes. The results show a wide divergence between the chromosomes of subsp. californicum and H. roshevitzii, and with their supposed derivatives in subsp. brachyantherum 4×. One of the three subgenomes of subsp. brachyantherum 6× is derived from subsp. gussoneanum with no genomic reorganisation (i.e., neither amplification nor loss of the repetitive DNA sequences analysed). It is generally accepted that subsp. brachyantherum 4× is the other progenitor of subsp. brachyantherum 6×, but the present results suggest this to be unlikely. The present findings thus show the cytogenetic diversity and genomic structure of H. brachyantherum, and reveal its complex evolutionary history, in which chromosomal diversification and allopolyploidy have played important roles.
... Likewise, H. chilense has been crossed with Secale species, obtaining hybrids with S. cereale... more ... Likewise, H. chilense has been crossed with Secale species, obtaining hybrids with S. cereale (FINCH & BENNET 1980, MARTIN & CUBERO 1981), S. africanum, S. montanum and S. vavilovii (FINCH & BENNETT 1980), and with 6x-Triticale (SCHRADER & POHLER 1985). ...
1991. Changes in expression of seed storage protein genes effected by chromosome 1D of wheat. Gen... more 1991. Changes in expression of seed storage protein genes effected by chromosome 1D of wheat. Genome, 34: 845-848. The effect of chromosome 1D of wheat on expression of seed storage protein genes was analyzed using a lR(1D) substitution line and derivatives of this line that had lost 1R (ID nullisomic). The results of one-and two-dimensional (two pH) electrophoresis suggested that nucleotide sequences located on ID were involved in induction and suppression of these protein genes and in posttranscriptional modification of polypeptides produced by the seed storage protein genes. The action attributed to ID appeared to affect only genes located on chromosome 6A.
Three repeated DNA sequences (rDNA 5S, 18S-5.8S-26S and telomeric repeats) were localised in the ... more Three repeated DNA sequences (rDNA 5S, 18S-5.8S-26S and telomeric repeats) were localised in the genomes ofLobelia brasiliensis andL. imperialis var.kanitzii (subg.Tupa), both with 2n = 28, by fluorescence in situ hybridization (FISH). The results were used to analyse the genomic relationship between the species. With probe pTa71, the karyotypes of these species showed only one NOR site. Probe pTa794, which
Simple Sequence Repeats (SSRs) are known to be scattered and present in high number in eukaryotic... more Simple Sequence Repeats (SSRs) are known to be scattered and present in high number in eukaryotic genomes. We demonstrate that dye-labeled oligodeoxyribonucleotides with repeated mono-, di-, tri, or tetranucleotide motifs (15-20 nucleotides in length) have an unexpected ability to recognize SSR target sequences in non-denatured chromosomes. The results show that all these probes are able to invade chromosomes, independent of the size of the repeat motif, their nucleotide sequence, or their ability to form alternative B-DNA structures such as triplex DNA. This novel and remarkable property of binding SSR oligonucleotides to duplex DNA targets permitted the development of a non-denaturing fluorescence in situ hybridization method that quickly and efficiently detects SSR-enriched chromosome regions in mitotic, meiotic, and polytene chromosome spreads of different model organisms. These results have implications for genome analysis and for investigating the roles of SSRs in chromosome structure and function.
The chromosome constitution and meiotic behaviour of 15 plants of F2 and BCI progenies from the h... more The chromosome constitution and meiotic behaviour of 15 plants of F2 and BCI progenies from the hybrid of 6xtriticale x T. turgidum (L.) ssp. turgidum cony. durum (Desf.) (Fl: 2n = 5x = 35; AABBR) were studied using Cbanding patterns of both wheat and rye chromosomes as identifying features. Patterns in PMCs at first metaphase permitted the individual identification of the chromosomes of nine of the wheat and all of rye in all plants. The external length and internal heterochromatin of chromosomes were analysed and found to influence the meiotic behaviour of each chromosome pair. The existence of effects of both arm length and amount and distribution of heterochromatin on pairing are assumed. The results seem to demonstrate that of the two mentioned factors the first is the most important.
Z-DNA forms transiently behind the active RNA polymerases, because of the mechanical torsional st... more Z-DNA forms transiently behind the active RNA polymerases, because of the mechanical torsional stress produced during transcription. In this paper, we explore the possibility that the distribution of Z-DNA stretches signals the sites related to nuclear transcription. To localize transcription, the in situ assay for active RNA polymerases, that allows the elongation of the already initiated transcripts but no initiation of new ones (run-on experiments), was carried out in isolated nuclei of Allium cepa L. root meristems. Both nucleolar and non-nucleolar sites appeared labelled. Nucleoli were most active in transcription than the multiple non-nucleolar foci altogether. In situ immunodetection of Z-DNA provided images that were comparable to those obtained after the run-on assay, with one exception: while Z-DNA and transcription sites were scattered throughout the whole nucleus, Z-DNA also accumulated in the nuclear periphery, where no transcription foci were detected in run-on assays. The peripheral Z-conformation signals might correspond to dsRNA segments present in the pre-mRNA in the process of their export to cytoplasm. The Z-containing structures nearly disappeared when non-nucleolar RNA polymerase II-dependent transcription had been previously abolished by the adenosine analogue DRB (5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole). This inhibition selectively decreased the amount of all nucleoplasmic Z structures. On the other hand, the inhibition of the nucleolar RNA polymerase I by cordycepin (3'-deoxyadenosine) prevented the presence of Z-DNA in nucleoli. We propose to use the in situ immunodetection of Z-DNA as a marker of the transcription level in both nucleolus and non-peripheral nucleoplasmic regions of nuclei. Co-detection of Z-DNA and of intermediate filament (IF) proteins, the major components of the nuclear matrix, was also carried out. The IFA antibody recognizes a conserved epitope essential for dimerization of the multiple IF proteins. They co-localized with most nucleolar Z-DNA, but not with the nucleoplasmic ones. In the nuclear periphery, the Z-positive signals were adjacent to the IF proteins constituting the lamina, though both signals did not often co-localize.
ABSTRACT Two intervarietal hybrids of common wheat, Triticum aestivum L., are meiotically analyze... more ABSTRACT Two intervarietal hybrids of common wheat, Triticum aestivum L., are meiotically analyzed using the C-banding staining method. The C-banding pattern of nine meiotic chromosomes (4A, 7A, and the seven of the B genome) permitted their unequivocal recognition at first metaphase plates. The pairing frequency of each B-genome chromosome arm was scored. Data on the pairing frequency of the arms, separately considered, are applied to calculate expected pairing of whole chromosomes and whole genomes. The application of mathematical models to predict the genome pairing using either equal or different frequencies per chromosome arm is discussed.Key words: meiotic analysis, Triticum aestivum L., C-banding.
SUMMARY - It is analyzed the primary structure of some of the high molecular weight (HMW) gluteni... more SUMMARY - It is analyzed the primary structure of some of the high molecular weight (HMW) glutenin genes and their degree of conservation in the genera Triticum, Secale and Aegilops. The primary structure presented information on the allelic composition and the quantities of the endosperm protein fractions (HMW, LMW and gliadins) in the doughs of different common wheat cultivars and lines in process of selection. Dough W is correlated with the quantity of HMW subunits present in the gluten and in particular with the quantity of type x HMW subunits. The gliadins and the LMW subunits appear to act as a "solvent" which modifies the rheological properties of the dough by interfering with the polymerization of the HMW subunits, or by altering the relative amounts of the different types of subunits available. The contribution of each type of subunit in the organization of the intermolecular links, the formation of the multiproteic aggregates, and the properties of the gluten, is...
El aborto es un tema muy espinoso y de complejo abordaje en el que fácilmente se pueden mezclar a... more El aborto es un tema muy espinoso y de complejo abordaje en el que fácilmente se pueden mezclar argumentos científicos o clínicos con otros de orden político o ideológico. De ahí la necesidad de partir de un razonamiento riguroso, honesto, y libre de todo tipo de prejuicio, error o engaño. Partiendo de esa premisa este trabajo se ha dividido en cuatro partes principales: una introducción en la que se hace una distinción entre un pensamiento riguroso y uno manipulador y las consecuencias que de ellos se derivan; una segunda parte en la que se hace una descripción del marco jurídico que definió las políticas de salud hoy vigentes en la materia; una tercera parte en la que se mide la evolución del aborto a lo largo de los últimos doce años, y finalmente, una última parte en la que se hace un análisis bioético del problema social que supone el aborto y de las consecuencias que se derivan de su práctica a nivel nacional.
TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik, 1984
The meiotic identification of nine pairs of chromosomes at metaphase I of meiosis of Triticum aes... more The meiotic identification of nine pairs of chromosomes at metaphase I of meiosis of Triticum aestivum (B genome, 4A and 7A) has been achieved using a Giemsa C-banding technique. As a result, the analysis of the pairing of each chromosome arm in disomic and monosomic intervarietal hybrids between 'Chinese Spring' and the Spanish cultivar 'Pané 247' could be carried out. Differences in the chiasmata frequencies per chromosome arm cannot be explained on the basis of relative arm lengths only. Possible effects of arm-to-arm heterochromatic differences on meiotic pairing are discussed.
SUMMARY - Most cereals have genomes with redundant genes arising from genome doubling or polyploi... more SUMMARY - Most cereals have genomes with redundant genes arising from genome doubling or polyploidy. Differences in the expression levels of duplicate genes have long been a subject of great interest since new genes can arise from redundant duplicates. Such analysis requires a technique that can discriminate between the expressions of two (or more) almost identical genes (homoeologues). In previous
Hordeum marinum is a species complex that includes the diploid subspecies marinum and both diploi... more Hordeum marinum is a species complex that includes the diploid subspecies marinum and both diploid and tetraploid forms of gussoneanum. Their relationships, the rank of the taxa and the origin of the polyploid forms remain points of debate. The present work reports a comparative karyotype analysis of six H. marinum accessions representing all taxa and cytotypes. Karyotypes were determined by analysing the chromosomal distribution of several tandemly repeated sequences, including the Triticeae cloned probes pTa71, pTa794, pAs1 and pSc119·2 and the simple sequence repeats (SSRs) (AG)10, (AAC)5, (AAG)5, (ACT)5 and (ATC)5. The identification of each chromosome pair in all subspecies and cytotypes is reported for the first time. Homologous relationships are also established. Wide karyotypic differences were detected within marinum accessions. Specific chromosomal markers characterized and differentiated the genomes of marinum and diploid gussoneanum. Two subgenomes were detected in the t...
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Papers by Nicolas Jouve