Characterization of the bacterial community involved in the bioflocculation process of wastewater organic matter in high loaded MBRs
Faust, L. ; Szendy, M. ; Plugge, C.M. ; Brink, P.F. van den; Temmink, H. ; Rijnaarts, H.H.M. - \ 2015
Applied Microbiology and Biotechnology 99 (2015)12. - ISSN 0175-7598 - p. 5327 - 5337.
solids retention time - 16s ribosomal-rna - gradient gel-electrophoresis - improved energy recovery - in-situ hybridization - activated-sludge - microbial community - membrane bioreactor - identification - stability
High-loaded membrane bioreactors (HL-MBRs), i.e., bioreactors equipped with a membrane for biomass retention and operated at extremely short sludge and hydraulic retention times, can concentrate sewage organic matter to facilitate subsequent energy and chemical recovery from these organics. Bioflocculation, accomplished by microorganisms that produce extracellular polymers, is a very important mechanism in these reactors. Bacterial diversity of the sludge and supernatant fraction of HL-MBRs operated at very short sludge retention times (0.125, 0.5, and 1 day) were determined using a PCR-denaturing gradient gel electrophoresis (DGGE) and clone library approach and compared to the diversity in sewage. Already at a sludge retention time (SRT) of 0.125 day, a distinct bacterial community developed compared to the community in sewage. Bioflocculation, however, was low and the majority of the bacteria, especially Arcobacter, were present in the supernatant fraction. Upon increasing SRT from 0.125 to 1 day, a much stronger bioflocculation was accompanied by an increased abundance of Bacteroidetes in the (solid) sludge fraction: 27.5 % at an SRT of 0.5 day and 46.4 % at an SRT of 1 day. Furthermore, cluster analysis of DGGE profiles revealed that the bacterial community structure in the sludge was different from that in the supernatant. To localize specific bacterial classes in the sludge flocs, fluorescence in situ hybridization (FISH) was carried out with three different bacterial probes. This showed that Betaproteobacteria formed clusters in the sludge flocs whereas Alphaproteobacteria and Gammaproteobacteria were mainly present as single cells
Introgression Browser: High throughput whole-genome SNP visualization
Aflitos, S.A. ; Sanchez Perez, G.F. ; Ridder, D. de; Fransz, P. ; Schranz, M.E. ; Jong, J.H.S.G.M. de; Peters, S.A. - \ 2015
The Plant Journal 82 (2015)1. - ISSN 0960-7412 - p. 174 - 182.
in-situ hybridization - alien chromosomes - recombination - tomato - markers - thaliana - potato - identification - organization - improvement
Breeding by introgressive hybridization is a pivotal strategy to broaden the genetic basis of crops. Usually, the desired traits are monitored in consecutive crossing generations by marker-assisted selection, but their analyses fail in chromosome regions where crossover recombinants are rare or not viable. Here, we present the Introgression Browser (IBROWSER), a bioinformatics tool aimed at visualizing introgressions at nucleotide or SNP accuracy. The software selects homozygous SNPs from Variant Call Format (VCF) information and filters out heterozygous SNPs, Multi-Nucleotide Polymorphisms (MNPs) and insertion-deletions (InDels). For data analysis IBROWSER makes use of sliding windows, but if needed it can generate any desired fragmentation pattern through General Feature Format (GFF) information. In an example of tomato (Solanum lycopersicum) accessions we visualize SNP patterns and elucidate both position and boundaries of the introgressions. We also show that our tool is capable of identifying alien DNA in a panel of the closely related S. pimpinellifolium by examining phylogenetic relationships of the introgressed segments in tomato. In a third example, we demonstrate the power of the IBROWSER in a panel of 597 Arabidopsis accessions, detecting the boundaries of a SNP-free region around a polymorphic 1.17 Mbp inverted segment on the short arm of chromosome 4. The architecture and functionality of IBROWSER makes the software appropriate for a broad set of analyses including SNP mining, genome structure analysis, and pedigree analysis. Its functionality, together with the capability to process large data sets and efficient visualization of sequence variation, makes IBROWSER a valuable breeding tool.
Characterization of B chromosomes in Lilium hybrids through GISH and FISH
Xie, S.L. ; Marasek-Ciolakowska, A. ; Ramanna, M.S. ; Arens, P.F.P. ; Visser, R.G.F. ; Tuyl, J.M. van - \ 2014
Plant Systematics and Evolution 300 (2014)8. - ISSN 0378-2697 - p. 1771 - 1777.
ribosomal-rna genes - in-situ hybridization - nuclear-dna amounts - plants - angiosperms - probe - recombination - isochromosome - misdivision - longiflorum
Supernumerary (B) chromosomes and small aberrant chromosomes were detected in Lilium hybrids and characterized through genomic in situ hybridization (GISH) and florescence in situ hybridization (FISH). Two small, supernumerary or B chromosomes were detected as extra chromosomes in a tetraploid plant derived from chromosome doubling of a hybrid (2n = 2x = 24) between a cultivar of the Longiflorum (L) and the Trumpet (T) group. When this tetraploid LLTT hybrid was crossed with a triploid LLO hybrid (O = Oriental), the B chromosome was transmitted to 73.4 % of the progenies. Based on GISH and FISH characterization, it was shown that the B chromosome consisted of two identical arms, with 5S rDNA hybridizing to the majority of it, which were flanked by normal telomeres, suggesting that this is an isochromosome. In another population, which is a backcross progeny between a F1 hybrid of Longiflorum x Asiatic (LA) and its Asiatic parent, the former produced functional 2n gametes which resulted in a triploid LAA progeny (2n = 3x = 36), in which three exceptional plants possessed 35 normal chromosomes and a small aberrant chromosome instead of the expected normal number of 36. In all three cases, the small aberrant chromosomes were isochromosomes which had obviously originated during the first backcross generation. These three chromosomes showed normal telomeres and mitosis. In addition, one of the new generated chromosomes possessed two 45S rDNA sites in the proximal positions. These new arisen isochromosomes were proposed to originate from centric breakage and fusion of two short arms of the missing chromosome in three genotypes, respectively, based on the comparison of arm lengths as well as rDNA loci. Their relevance to the origin of Bs is discussed.
Chromosomal organizations of major repeat families on potato (Solanum tuberosum) and further exploring in its sequenced genome
Tang, X. ; Datema, E. ; Olortegui Guzman, M.C. ; Boer, J.M. de; Eck, H.J. van; Bachem, C.W.B. ; Visser, R.G.F. ; Jong, H. de - \ 2014
Molecular Genetics and Genomics 289 (2014)6. - ISSN 1617-4615 - p. 1307 - 1319.
ty1-copia group retrotransposons - species-specific sequences - repeated dna-sequences - in-situ hybridization - somatic hybrids - lycopersicon-esculentum - satellite repeat - repetitive dna - transposable elements - metaphase chromosomes
One of the most powerful technologies in unraveling the organization of a eukaryotic plant genome is high-resolution Fluorescent in situ hybridization of repeats and single copy DNA sequences on pachytene chromosomes. This technology allows the integration of physical mapping information with chromosomal positions, including centromeres, telomeres, nucleolar-organizing region, and euchromatin and heterochromatin. In this report, we established chromosomal positions of different repeat fractions of the potato genomic DNA (Cot100, Cot500 and Cot1000) on the chromosomes. We also analysed various repeat elements that are unique to potato including the moderately repetitive P5 and REP2 elements, where the REP2 is part of a larger Gypsy-type LTR retrotransposon and cover most chromosome regions, with some brighter fluorescing spots in the heterochromatin. The most abundant tandem repeat is the potato genomic repeat 1 that covers subtelomeric regions of most chromosome arms. Extensive multiple alignments of these repetitive sequences in the assembled RH89-039-16 potato BACs and the draft assembly of the DM1-3 516 R44 genome shed light on the conservation of these repeats within the potato genome. The consensus sequences thus obtained revealed the native complete transposable elements from which they were derived
GISH analyses of backcross progenies of two Lilium species hybrids and their relevance to breeding
Luo, J.R. ; Ramanna, M.S. ; Arens, P. ; Niu, L.X. ; Tuyl, J.M. van - \ 2012
Journal of Horticultural Science and Biotechnology 87 (2012)6. - ISSN 1462-0316 - p. 654 - 660.
in-situ hybridization - x asiatic hybrids - sexual polyploidization - interspecific hybridization - intergenomic recombination - homoeologous recombination - bc2 progenies - lily hybrids - longiflorum - introgression
A total of 21 BC1 progenies of Oriental × Trumpet (OT) Lilium hybrids, nine BC2 progenies of OT hybrids, and three BC1 progenies of Martagon × Asiatic (MA) hybrids were analysed using genomic in situ hybridisation (GISH). Of the BC1 progenies of the OT hybrids, 15 were euploid (14 triploids, 2n = 3x = 36; 1 diploid, 2n = 2x = 24), and six were aneuploid. The triploid progenies had resulted from 2n eggs due to first division restitution (FDR). The aneuploid progenies had resulted from viable aneuploid gametes. All the BC2 progenies of triploid (2n = 3x = 36) OOT hybrids were aneuploid. Chromosome numbers in the BC2 progenies varied from 25 to 29.Two out of three BC1 progenies of the MA hybrids were aneuploids, with 35 and 32 chromosomes, respectively. Another was a triploid (2n = 3x = 36) which had resulted from 2n eggs due to indeterminate meiotic restitution (IMR). GISH revealed extensive intergenomic recombination in the BC1 progenies of OT and MA hybrids. A large number of Trumpet or Martagon chromosome segments were transmitted to the BC1 progenies from the F1 OT or MA hybrids. However, very few Trumpet chromosome segments were transmitted from the triploid BC1 parent to the BC2 progenies. These results indicate that it should be possible to select cultivars which possessed both traits from these BC1 progenies and they would be fertile.
Assessment of intergenomic recombination through GISH analysis of F1, BC1 and BC2 progenies of Tulipa gesneriana and T. fosteriana
Marasek Ciolakowska, A.R. ; He, H. ; Bijman, P.J.J. ; Ramanna, M.S. ; Arens, P. ; Tuyl, J.M. van - \ 2012
Plant Systematics and Evolution 298 (2012)5. - ISSN 0378-2697 - p. 887 - 899.
in-situ hybridization - sexual polyploidization - species relationships - chromosome variation - herbaceous plants - lilium hybrids - garden tulips - clusiana dc - cultivars - liliaceae
Using 23 F1 hybrids, 14 BC1 and 32 BC2 progenies, the genome composition of Darwin hybrid tulips was analysed through genomic in situ hybridisation (GISH) of somatic chromosomes. All plants were diploids (2n = 2x = 24) with the exception of one tetraploid BC1 (2n = 4x = 48) and one aneuploid BC2 (2n = 2x + 1 = 25) hybrid. Morphometric analysis in F1 hybrids revealed a difference in the total length of chromosomes representing genomes of T. gesneriana and T. fosteriana, where the percentage of each genome equaled 55.18 ± 0.8 and 44.92 ± 0.6% respectively. GISH distinguished chromosomes from both parent genomes although there was a lack of consistent chromosome labelling in some cases. In both T. gesneriana and T. fosteriana chromosomes some segments of heterochromatin in the telomeric and intercalary regions exhibited a higher intensity of fluorescence. In situ hybridisation with 5S rDNA and 45S rDNA probes to metaphase chromosomes of F1 hybrids showed that these regions are rich in rDNA. A notable feature was that, despite genome differences, there was a considerable amount of intergenomic recombination between the parental chromosomes of the two species as estimated in both BC1 and BC2 offspring. The number of recombinant chromosomes ranged from 3 to 8 in BC1 and from 1 to 7 in BC2 progenies. All recombinant chromosomes possessed mostly a single recombinant segment derived from either a single crossover event or in a few cases double crossover events. This explains the fact that, unlike the situation in most F1 hybrids of other plant species, certain genotypes of Darwin hybrid tulips behave like normal diploid plants producing haploid gametes and give rise to mostly diploid sporophytes.
Role of syntrophic microbial communities in high-rate methanogenic bioreactors
Stams, A.J.M. ; Sousa, D.Z. ; Kleerebezem, R. ; Plugge, C.M. - \ 2012
Water Science and Technology 66 (2012)2. - ISSN 0273-1223 - p. 352 - 362.
chain fatty-acids - propionate-oxidizing bacterium - in-situ hybridization - interspecies electron-transfer - spore-forming bacterium - wolfei subsp saponavida - municipal solid-waste - sp-nov. - gen. nov. - anaerobic treatment
Anaerobic purification is a cost-effective way to treat high strength industrial wastewater. Through anaerobic treatment of wastewaters energy is conserved as methane, and less sludge is produced. For high-rate methanogenesis compact syntrophic communities of fatty acid-degrading bacteria and methanogenic archaea are essential. Here, we describe the microbiology of syntrophic communities in methanogenic reactor sludges and provide information on which microbiological factors are essential to obtain high volumetric methane production rates. Fatty-acid degrading bacteria have been isolated from bioreactor sludges, but also from other sources such as freshwater sediments. Despite the important role that fatty acid-degrading bacteria play in high-rate methanogenic bioreactors, their relative numbers are generally low. This finding indicates that the microbial community composition can be further optimized to achieve even higher rates
The banana (Musa acuminata) genome and the evolution of monocotyledonous plants
Hont, A. D'; Denoeud, F. ; Aury, J.M. ; Kema, G.H.J. ; Dita Rodriguez, M.A. ; Waalwijk, C. - \ 2012
Nature 488 (2012). - ISSN 0028-0836 - p. 213 - 217.
in-situ hybridization - sequence count data - dna-sequences - differential expression - maximum-likelihood - gene - rice - diversification - identification - resource
Bananas (Musa spp.), including dessert and cooking types, are giant perennial monocotyledonous herbs of the order Zingiberales, a sister group to the well-studied Poales, which include cereals. Bananas are vital for food security in many tropical and subtropical countries and the most popular fruit in industrialized countries1. The Musa domestication process started some 7,000 years ago in Southeast Asia. It involved hybridizations between diverse species and subspecies, fostered by human migrations2, and selection of diploid and triploid seedless, parthenocarpic hybrids thereafter widely dispersed by vegetative propagation. Half of the current production relies on somaclones derived from a single triploid genotype (Cavendish)1. Pests and diseases have gradually become adapted, representing an imminent danger for global banana production3, 4. Here we describe the draft sequence of the 523-megabase genome of a Musa acuminata doubled-haploid genotype, providing a crucial stepping-stone for genetic improvement of banana. We detected three rounds of whole-genome duplications in the Musa lineage, independently of those previously described in the Poales lineage and the one we detected in the Arecales lineage. This first monocotyledon high-continuity whole-genome sequence reported outside Poales represents an essential bridge for comparative genome analysis in plants. As such, it clarifies commelinid-monocotyledon phylogenetic relationships, reveals Poaceae-specific features and has led to the discovery of conserved non-coding sequences predating monocotyledon–eudicotyledon divergence
Microbial community structure elucidates performance of Glyceria maxima plant microbial fuel cell
Timmers, R.A. ; Rothballer, M. ; Strik, D.P.B.T.B. ; Engel, M. ; Schulz, M. ; Hartmann, A. ; Hamelers, H.V.M. ; Buisman, C.J.N. - \ 2012
Applied Microbiology and Biotechnology 94 (2012)2. - ISSN 0175-7598 - p. 537 - 548.
targeted oligonucleotide probes - iron-reducing bacteria - in-situ hybridization - electricity-generation - fe(iii)-reducing bacterium - shewanella-putrefaciens - activated-sludge - soil bacteria - rice plants - human feces
The plant microbial fuel cell (PMFC) is a technology in which living plant roots provide electron donor, via rhizodeposition, to a mixed microbial community to generate electricity in a microbial fuel cell. Analysis and localisation of the microbial community is necessary for gaining insight into the competition for electron donor in a PMFC. This paper characterises the anode-rhizosphere bacterial community of a Glyceria maxima (reed mannagrass) PMFC. Electrochemically active bacteria (EAB) were located on the root surfaces, but they were more abundant colonising the graphite granular electrode. Anaerobic cellulolytic bacteria dominated the area where most of the EAB were found, indicating that the current was probably generated via the hydrolysis of cellulose. Due to the presence of oxygen and nitrate, short-chain fatty acid-utilising denitrifiers were the major competitors for the electron donor. Acetate-utilising methanogens played a minor role in the competition for electron donor, probably due to the availability of graphite granules as electron acceptors.
Genetic characterization of a reciprocal translocation present in a widely grown barley variety
Farré, A. ; Cuadrado, A. ; Lacasa-Benito, I. ; Cistué, L. ; Schubert, I. ; Comadran, J. ; Jansen, J. ; Romagosa, I. - \ 2012
Molecular Breeding 30 (2012)2. - ISSN 1380-3743 - p. 1109 - 1119.
in-situ hybridization - nonhomologous chromosomes - genome duplication - hordeum-vulgare - breakpoints - map - localization - interchange - reduction - markers
Artificially induced translocation stocks have been used to physically map the barley genome; however, natural translocations are extremely uncommon in cultivated genotypes. Albacete is a barley variety widely grown in recent decades in Spain and carrying a reciprocal translocation which obviously does not affect its agronomical fitness. This translocation has been characterized by a combination of cytological and molecular genetic approaches. Firstly, recombination frequencies between markers on chromosomes 1H and 3H were estimated to determine the boundaries of the reciprocal interchange. Secondly, 1H-3H wheat barley telosome addition lines were used to assign selected markers to chromosome arms. Thirdly, fluorescence in situ hybridization (FISH) with rDNA probes (5S and 18S-5.8S-26S) and microsatellite probes [(ACT)5, (AAG)5 and (CAG)5] was used to determine the locations of the translocation breakpoints more precisely. Fourthly, fine-mapping of the regions around the translocation breakpoints was used to increase the marker density for comparative genomics. The results obtained in this study indicate that the translocation is quite large with breakpoints located on the long arms of chromosomes 1H and 3H, between the pericentromeric (AAG)5 bands and above the (ACT)5 interstitial distal bands, resulting in the reciprocal translocation 1HS.1HL-3HL and 3HS.3HL-1HL. The gene content around the translocation breakpoints could be inferred from syntenic relationships observed among different species from the grass family Poaceae (rice, Sorghum and Brachypodium) and was estimated at approximately 1,100 and 710 gene models for 1H and 3H, respectively. Duplicated segments between chromosomes Os01 and Os05 in rice derived from ancestral duplications within the grass family overlap with the translocation breakpoints on chromosomes 1H and 3H in the barley variety Albacete.
Genome constitution of Narcissus variety, 'Tete-a-Tete', analysed through GISH and NBS profiling
Wu, H. ; Ramanna, M.S. ; Arens, P. ; Tuyl, J.M. van - \ 2011
Euphytica 181 (2011)2. - ISSN 0014-2336 - p. 285 - 292.
in-situ hybridization - homoeologous recombination - hybrid - origin - chromosomes - lilium - dna
The Narcissus variety, ‘Tête-à-Tête’, has been the most popular variety since 1949, and a well known allotriploid (2n = 3x = 24 + B) of spontaneous origin. Because the identity of one of the parents of this variety was uncertain, the genome constitution of ‘Tête-à-Tête’ was investigated by using genomic in situ hybridization (GISH) and NBS profiling. Both of these techniques confirmed that two different species of Narcissus, viz., N. tazetta (2n = 2x = 20) and N. cyclamineus (2n = 2x = 14) are the parents. GISH clearly identified 10 chromosomes of N. tazetta and 14 chromosomes of N. cyclamineus, the former has contributed one and the latter has contributed two genomes. One B chromosome was identically labelled as those of N. cyclamineus indicating the affinity of the special chromosome to this species. Due to its male and female sterility ‘Tête-à-Tête’ is unsuitable as a parent for utilizing in further breeding, it might be possible to re-synthesize a ‘Tête-à-Tête’ like variety using the now known parents and the original pathway
Advances in research on the prenatal development of skeletal muscle in animals in relation to the quality of muscle-based food. I - Regulation of myogenesis and environmental impact
Rehfeldt, C. ; Pas, M.F.W. te; Wimmers, K. ; Brameld, J.M. ; Nissen, P.M. ; Berri, C. ; Valente, L.M.P. ; Power, D.M. ; Picard, B. ; Stickland, N.C. ; Oksbjerg, N. - \ 2011
Animal 5 (2011)5. - ISSN 1751-7311 - p. 703 - 717.
trout oncorhynchus-mykiss - receptor signal-transduction - seabream pagellus-bogaraveo - in-situ hybridization - gilthead sea bream - salmon salmo-salar - rainbow-trout - birth-weight - meat quality - igf-i
Skeletal muscle development in vertebrates – also termed myogenesis – is a highly integrated process. Evidence to date indicates that the processes are very similar across mammals, poultry and fish, although the timings of the various steps differ considerably. Myogenesis is regulated by the myogenic regulatory factors and consists of two to three distinct phases when different fibre populations appear. The critical times when myogenesis is prone to hormonal or environmental influences depend largely on the developmental stage. One of the main mechanisms for both genetic and environmental effects on muscle fibre development is via the direct action of the growth hormone–insulin-like growth factor (GH–IGF) axis. In mammals and poultry, postnatal growth and function of muscles relate mainly to the hypertrophy of the fibres formed during myogenesis and to their fibre-type composition in terms of metabolic and contractile properties, whereas in fish hyperplasia still plays a major role. Candidate genes that are important in skeletal muscle development, for instance, encode for IGFs and IGF-binding proteins, myosin heavy chain isoforms, troponin T, myosin light chain and others have been identified. In mammals, nutritional supply in utero affects myogenesis and the GH–IGF axis may have an indirect action through the partitioning of nutrients towards the gravid uterus. Impaired myogenesis resulting in low skeletal myofibre numbers is considered one of the main reasons for negative long-term consequences of intrauterine growth retardation. Severe undernutrition in utero due to natural variation in litter or twin-bearing species or insufficient maternal nutrient supply may impair myogenesis and adversely affect carcass quality later in terms of reduced lean and increased fat deposition in the progeny. On the other hand, increases in maternal feed intake above standard requirement seem to have no beneficial effects on the growth of the progeny with myogenesis not or only slightly affected. Initial studies on low and high maternal protein feeding are published. Although there are only a few studies, first results also reveal an influence of nutrition on skeletal muscle development in fish and poultry. Finally, environmental temperature has been identified as a critical factor for growth and development of skeletal muscle in both fish and poultry.
Comparison of the chromosome maps around a resistance hot spot on chromosome 5 of potato and tomato using BAC-FISH painting
Achenbach, U.C. ; Tang, X.M. ; Ballvora, A. ; Jong, H. de; Gebhardt, C. - \ 2010
Genome 53 (2010)2. - ISSN 0831-2796 - p. 103 - 110.
in-situ hybridization - zea-mays l. - broad-spectrum resistance - globodera-pallida - phytophthora-infestans - pachytene chromosomes - gene-cluster - metaphase chromosomes - arabidopsis-thaliana - solanum-spegazzinii
Potato chromosome 5 harbours numerous genes for important qualitative and quantitative traits, such as resistance to the root cyst nematode Globodera pallida and the late blight fungus, Phytophthora infestans. The genes make up part of a “hot spot” for resistances to various pathogens covering a genetic map length of 3 cM between markers GP21 and GP179. We established the physical size and position of this region on chromosome 5 in potato and tomato using fluorescence in situ hybridization (FISH) on pachytene chromosomes. Five potato bacterial artificial chromosome (BAC) clones with the genetically anchored markers GP21, R1-contig (proximal end), CosA, GP179, and StPto were selected, labeled with different fluorophores, and hybridized in a five-colour FISH experiment. Our results showed the location of the BAC clones in the middle of the long arm of chromosome 5 in both potato and tomato. Based on chromosome measurements, we estimate the physical size of the GP21–GP179 interval at 0.85 Mb and 1.2 Mb in potato and tomato, respectively. The GP21–GP179 interval is part of a genome segment known to have inverted map positions between potato and tomato.
An assessment of chromosomal rearrangements in neopolyploids of Lilium hybrids
Xie, S.L. ; Khan, N. ; Ramanna, M.S. ; Niu, L.X. ; Marasek Ciolakowska, A.R. ; Arens, P. ; Tuyl, J.M. van - \ 2010
Genome 53 (2010)6. - ISSN 0831-2796 - p. 439 - 446.
in-situ hybridization - x asiatic hybrids - brassica-napus - intergenomic recombination - homeologous recombination - flowering plants - gish analysis - genome - longiflorum - polyploids
Two types of newly induced polyploids (neopolyploids) of Lilium hybrids were monitored for the occurrence of chromosomal rearrangements through genomic in situ hybridization (GISH) technique. One of the populations was obtained through crossing an allotriploid Longiflorum x Oriental hybrid (LLO) with an allotetraploid Longiflorum x Trumpet hybrid (LLTT), both of which were derived from somatic chromosome doubling. The other type of allopolyploid population was derived from meiotic chromosome doubling in which numerically unreduced (2n) gametes from two different interspecific hybrids. namely. Longiflorum x Asiatic (LA) and Oriental x Asiatic (OA). were used to get backcross progeny with the Asiatic parents. GISH clearly discriminated the three constituent genomes (L, T, and O) in the complements of the progeny obtained from mitotic chromosome doubling. A total of 26 individuals were analyzed from this population and there was no evidence of chromosomal rearrangements. However, in the case of meiotically doubled allopolyploid progeny, considerable frequencies of chromosomal rearrangements were observed through GISH. The so-called chromosomal rearrangements in meiotic polyploids are the result of homoeologous recombination rather than translocations. Furthermore, evidence for the occurrence of meiotic recombination in the LA hybrids has been confirmed with GISH on meiotic chromosomes. Thus, there was evidence that neopolyploids of Lilium hybrids did not possess any noticeable chromosome rearrangements
Molecular cytogenetic mapping of Cucumis sativus and C. melo using highly repetitive DNA sequences
Koo, D.H. ; Nam, Y.W. ; Choi, D. ; Bang, J.W. ; Jong, J.H.S.G.M. de; Hur, Y. - \ 2010
Chromosome Research 18 (2010). - ISSN 0967-3849 - p. 325 - 336.
in-situ hybridization - alpha-satellite dna - arabidopsis-thaliana - robertsonian translocations - mitochondrial genome - tandem repeats - evolution - organization - centromere - chromosomes
Chromosomes often serve as one of the most important molecular aspects of studying the evolution of species. Indeed, most of the crucial mutations that led to differentiation of species during the evolution have occurred at the chromosomal level. Furthermore, the analysis of pachytene chromosomes appears to be an invaluable tool for the study of evolution due to its effectiveness in chromosome identification and precise physical gene mapping. By applying fluorescence in situ hybridization of 45S rDNA and CsCent1 probes to cucumber pachytene chromosomes, here, we demonstrate that cucumber chromosomes 1 and 2 may have evolved from fusions of ancestral karyotype with chromosome number n= 12. This conclusion is further supported by the centromeric sequence similarity between cucumber and melon, which suggests that these sequences evolved from a common ancestor. It may be after or during speciation that these sequences were specifically amplified, after which they diverged and specific sequence variants were homogenized. Additionally, a structural change on the centromeric region of cucumber chromosome 4 was revealed by fiber-FISH using the mitochondrial-related repetitive sequences, BAC-E38 and CsCent1. These showed the former sequences being integrated into the latter in multiple regions. The data presented here are useful resources for comparative genomics and cytogenetics of Cucumis and, in particular, the ongoing genome sequencing project of cucumber
Alternative microbial methods: An overview and selection criteria.
Jasson, V. ; Jacxsens, L. ; Luning, P.A. ; Rajkovic, A. ; Uyttendaele, M. - \ 2010
Food Microbiology 27 (2010)6. - ISSN 0740-0020 - p. 710 - 730.
escherichia-coli o157-h7 - real-time pcr - latex agglutination tests - atp-bioluminescence method - in-situ hybridization - plate-count method - listeria-monocytogenes - rapid detection - raw-milk - flow-cytometry
This study provides an overview and criteria for the selection of a method, other than the reference method, for microbial analysis of foods. In a first part an overview of the general characteristics of rapid methods available, both for enumeration and detection, is given with reference to relevant bibliography. Perspectives on future development and the potential of the rapid method for routine application in food diagnostics are discussed. As various alternative “rapid” methods in different formats are available on the market, it can be very difficult for a food business operator or for a control authority to select the most appropriate method which fits its purpose. Validation of a method by a third party, according to international accepted protocol based upon ISO 16140, may increase the confidence in the performance of a method. A list of at the moment validated methods for enumeration of both utility indicators (aerobic plate count) and hygiene indicators (Enterobacteriaceae, Escherichia coli, coagulase positive Staphylococcus) as well as for detection of the four major pathogens (Salmonella spp., Listeria monocytogenes, E. coli O157 and Campylobacter spp.) is included with reference to relevant websites to check for updates. In a second part of this study, selection criteria are introduced to underpin the choice of the appropriate method(s) for a defined application. The selection criteria link the definition of the context in which the user of the method functions – and thus the prospective use of the microbial test results – with the technical information on the method and its operational requirements and sustainability. The selection criteria can help the end user of the method to obtain a systematic insight into all relevant factors to be taken into account for selection of a method for microbial analysis
FISH applications for genomics and plant breeding strategies in tomato and other Solanaceous crops
Szinay, D. ; Bai, Y. ; Visser, R.G.F. ; Jong, J.H. de - \ 2010
Cytogenetic and Genome Research 129 (2010)1-3. - ISSN 1424-8581 - p. 199 - 210.
in-situ hybridization - high-resolution fish - extended dna fibers - lycopersicon-esculentum - recombination nodules - heterochromatic genes - chromosome identification - synaptonemal complexes - pachytene chromosomes - sequence-analysis
This paper describes the use of advanced fluorescence in situ hybridization (FISH) technologies for genomics and breeding of tomato and related Solanum species. The first part deals with the major determinants of FISH technology: (1) spatial resolution, which depends on the diffraction limit of the microscope and the type of chromosome, chromatin or isolated DNA fibres as target for the hybridisation; (2) the detection sensitivity, which is limited by the sensitivity and dynamic range of the CCD camera and the quality of the microscope, and the amplification system of the weak signals from tiny probe molecules; (3) simultaneous detection of multiple probes labelled directly or indirectly with up to 5 different fluorophores, whether or not in different combinations and/or mixed at different ratios. The power and usability of such multicolour FISH is indispensable when large numbers of bacterial artificial chromosomes (BACs) or other vectors with genomic DNA are available. Mapping of multiple BACs on chromosomes are powerful instruments confirming their assumed genetic position, whereas pooled BACs for a given chromosome arm will reveal the gaps between the BACs or derived contigs of their physical maps. Tandem and dispersed repeats, which are abundant in the genomes of most species, can be analysed in repeat bar coding FISH, showing the major blocks of repeats in heterochromatin and euchromatin areas. Repeat-rich areas of the chromosomes can also be demonstrated by hybridisation of probed Cot fractions of sheared genomic DNA; a powerful method to elucidate the heterochromatin domains for genomic studies. In addition, unlabelled Cot DNA, as blocking agent in BAC-FISH painting, suppresses repetitive sequences from the BACs to hybridise on the chromosomes. Cross-species BAC-FISH painting with labelled probes from tomato and potato BACs and hybridised on the chromosomes of related species, under appropriate conditions, is a powerful instrument to demonstrate chromosomal rearrangements, including inversions and translocations. The technology not only supports phylogenetic studies between the taxa under study but can also be helpful in breeding programs with crops containing introgressed regions from related species when linkage drag or meiotic pairing disturbances between the homoeologues are assumed. In the next steps in comparative genomics, we now can detect smaller chromosomal and DNA rearrangements, diminutions and amplifications of repeats and changes of the epigenetic status of introgressed regions
GISH Analysis of Subsequent Progeny Crossed with 2n-gametes of F-1 Oriental-Asiatic Interspecific Hybrid in Lily
Chung, M.Y. ; Chung, J.D. ; Tuyl, J.M. van; Lim, K.B. - \ 2009
Korean Journal of Horticultural Science and Technology 27 (2009)4. - ISSN 1226-8763 - p. 649 - 656.
in-situ hybridization - alfalfa medicago-sativa - 2n pollen formation - homoeologous recombination - lilium-longiflorum - diploid alfalfa - distant hybrid - fertility - polyploidization - alstroemeria
This study was carried out to analyze the chromosome composition and homoeologous recombination of subsequent progenies, derived from cross of Oriental-Asiatic (OA) interspecific hybrid producing 2n-gametes, by GISH technique. GISH analysis of F-1 OA interspecific hybrid '2nOA-001', which produced 2n-gametes, showed 24 chromosomes consisting of 12 chromosomes from Oriental hybrid and 12 chromosomes from Asiatic hybrid. The subsequent backcross progeny of A x OA (2x-2x) crosses produced diploid plants with all Asiatic chromosomes (2x=24) and some triploids (O=12 and A=24), and one aneuploid 2n=35 (O=11 and A=24). Seven plants were produced in the line of 'AOA-006', among them, it was confirmed that three plants were homoeologous recombination with I or 2 breakpoints at short- or long-arm randomly. In case of 0 x OA (2x-2x) crosses, all plants showed 2n=24 chromosomes that originated from Oriental hybrid and no homoeologous recombination. In case of OA x A (2x-2x) crosses, triploid plants (O=12, A=24) were formed without any homoeologous recombination. All progenies were tetraploid plants in the cross of OA x 00 (2x-4x) showing 36 chromosomes that originated from Oriental hybrid and 12 chromosomes originated from Asiatic hybrid, without any homoeologous recombinations.
Assignment of genetic linkage maps to diploid Solanum tuberosum pachytene chromosomes by BAC-FISH technology
Tang, X. ; Boer, J.M. de; Eck, H.J. van; Bachem, C.W.B. ; Visser, R.G.F. ; Jong, J.H. de - \ 2009
Chromosome Research 17 (2009)7. - ISSN 0967-3849 - p. 899 - 915.
in-situ hybridization - bacterial artificial chromosomes - tomato lycopersicon-esculentum - cytogenetic dna markers - metaphase chromosomes - pericentromeric heterochromatin - arabidopsis-thaliana - meiotic pachytene - rice centromere - aflp markers
A cytogenetic map has been developed for diploid potato (Solanum tuberosum), in which the arms of the 12 potato bivalents can be identified in pachytene complements using multicolor fluorescence in situ hybridization (FISH) with a set of 60 genetically anchored bacterial artificial chromosome (BAC) clones from the RHPOTKEY BAC library. This diagnostic set of selected BACs (five per chromosome) hybridizes to euchromatic regions and corresponds to well-defined loci in the ultradense genetic map, and with these probes a new detailed and reliable pachytene karyotype could be established. Chromosome size has been estimated both from microscopic length measurements and from 4',6-diamidino-2-phenylindole fluorescence-based DNA content measurements. In both approaches, chromosome 1 is the largest (100–115 Mb) and chromosome 11 the smallest (49–53 Mb). Detailed measurements of mega-base-pair to micrometer ratios have been obtained for chromosome 5, with average values of 1.07 Mb/µm for euchromatin and 3.67 Mb/µm for heterochromatin. In addition, our FISH results helped to solve two discrepancies in the potato genetic map related to chromosomes 8 and 12. Finally, we discuss the significance of the potato cytogenetic map for extended FISH studies in potato and related Solanaceae, which will be especially beneficial for the potato genome-sequencing project
Construction of chromosomal recombination maps of three genomes of lilies (Lilium) based on GISH analysis.
Nadeem Khan, M. ; Shujun Zhou, ; Barba Gonzalez, R. ; Ramanna, M.S. ; Visser, R.G.F. ; Tuyl, J.M. van - \ 2009
Genome 52 (2009)3. - ISSN 0831-2796 - p. 238 - 251.
in-situ hybridization - genetic-linkage maps - nuclear-dna amounts - x asiatic hybrids - translocation breakpoints - intergenomic recombination - homoeologous recombination - sexual polyploidization - interspecific hybrids - arabidopsis-thaliana
Chromosomal recombination maps were constructed for three genomes of lily (Lilium) using GISH analyses. For this purpose, the backcross (BC) progenies of two diploid (2n = 2x = 24) interspecific hybrids of lily, viz. Longiflorum × Asiatic (LA) and Oriental × Asiatic (OA), were used. Mostly the BC progenies of LA hybrids consisted of both triploid (2n = 3x = 36) and diploid (2n = 2x = 24) with some aneuploid genotypes and those of OA hybrids consisted of triploid (2n = 3x = 36) and some aneuploid genotypes. In all cases, it was possible to identify the homoeologous recombinant chromosomes as well as accurately count the number of crossover points, which are called ¿recombination sites¿. Recombination sites were estimated in the BC progeny of 71 LA and 41 OA genotypes. In the case of BC progenies of LA hybrids, 248 recombination sites were cytologically localized on 12 different chromosomes of each genome (i.e., L and A). Similarly, 116 recombinant sites were localized on the 12 chromosomes each from the BC progenies of OA hybrids (O and A genomes). Cytological maps were constructed on the basis of the percentages of distances (micrometres) of the recombination sites from the centromeres. Since an Asiatic parent was involved in both hybrids, viz. LA and OA, two maps were constructed for the A genome that were indicated as Asiatic (L) and Asiatic (O). The other two maps were Longiflorum (A) and Oriental (A). Remarkably, the recombination sites were highly unevenly distributed among the different chromosomes of all four maps. Because the recombination sites can be unequivocally identified through GISH, they serve as reliable landmarks and pave the way for assigning molecular markers or desirable genes to chromosomes of Lilium and also monitor introgression of alien segments