A Bacterial Artificial Chromosome Contig Spanning the Major Domestication Locus Q in Wheat and Identification of a Candidate Gene

Abstract The Q locus played a major role in the domestication of wheat because it confers the free-threshing character and influences many other agronomically important traits. We constructed a physical contig spanning the Q locus using a Triticum monococcum BAC library. Three chromosome walking steps were performed by complete sequencing of BACs and identification of low-copy markers through similarity searches of database sequences. The BAC contig spans a physical distance of ∼300 kb corresponding to a genetic distance of 0.9 cM. The physical map of T. monococcum had perfect colinearity with the genetic map of wheat chromosome arm 5AL. Recombination data in conjunction with analysis of fast neutron deletions confirmed that the contig spanned the Q locus. The Q gene was narrowed to a 100-kb segment, which contains an APETALA2 (AP2)-like gene that cosegregates with Q. AP2 is known to play a major role in controlling floral homeotic gene expression and thus is an excellent candidate for Q.

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A4Mb High Resolution BAC Contig on Bovine Chromosome1q12and Comparative Analysis With Human Chromosome21q22

Comparative and Functional Genomics ◽

10.1002/cfg.476 ◽

2005 ◽

Vol 6 (4) ◽

pp. 194-203 ◽

Cited By ~ 8

Author(s):

Cord Drögemüller ◽

Anne Wöhlke ◽

Tosso Leeb ◽

Ottmar Distl

Keyword(s):

Positional Cloning ◽

Physical Map ◽

Dominant Gene ◽

Bac Library ◽

Exact Order ◽

Bac Contig ◽

Nucleotide Sequence Data ◽

New Genes ◽

Human Genes ◽

First Time

The bovine RPCI-42 BAC library was screened to construct a sequence-ready ~4 Mb single contig of 92 BAC clones on BTA 1q12. The contig covers the region between the genesKRTAP8P1andCLIC6. This genomic segment in cattle is of special interest as it contains the dominant gene responsible for the hornless or polled phenotype in cattle. The construction of the BAC contig was initiated by screening the bovine BAC library with heterologous cDNA probes derived from 12 human genes of the syntenic region on HSA 21q22. Contig building was facilitated by BAC end sequencing and chromosome walking. During the construction of the contig, 165 BAC end sequences and 109 single-copy STS markers were generated. For comparative mapping of 25 HSA 21q22 genes, genomic PCR primers were designed from bovine EST sequences and the gene-associated STSs mapped on the contig. Furthermore, bovine BAC end sequence comparisons against the human genome sequence revealed significant matches to HSA 21q22 and allowed thein silicomapping of two new genes in cattle. In total, 31 orthologues of human genes located on HSA 21q22 were directly mapped within the bovine BAC contig, of which 16 genes have been cloned and mapped for the first time in cattle. In contrast to the existing comparative bovine–human RH maps of this region, these results provide a better alignment and reveal a completely conserved gene order in this 4 Mb segment between cattle, human and mouse. The mapping of known polled linked BTA 1q12 microsatellite markers allowed the integration of the physical contig map with existing linkage maps of this region and also determined the exact order of these markers for the first time. Our physical map and transcript map may be useful for positional cloning of the putative polled gene in cattle. The nucleotide sequence data reported in this paper have been submitted to EMBL and have been assigned Accession Numbers AJ698510–AJ698674.

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Physical mapping of resistant and susceptible soybean genomes near the soybean cyst nematode resistance gene Rhg4

Genome ◽

10.1139/g01-109 ◽

2001 ◽

Vol 44 (6) ◽

pp. 1057-1064 ◽

Cited By ~ 4

Author(s):

K S Lewers ◽

S D Nilmalgoda ◽

A L Warner ◽

H T Knap ◽

B F Matthews

Keyword(s):

Resistance Gene ◽

Soybean Cyst Nematode ◽

Bac Library ◽

Artificial Chromosome ◽

Nematode Resistance ◽

Physical Distance ◽

Cyst Nematode ◽

Nematode Resistance Gene ◽

Glycine Max L ◽

Soybean Cyst Nematode Resistance

The soybean cyst nematode (SCN), Heterodera glycines Ichinohe, is the foremost pest of soybean (Glycine max L. Merr.). The rhg1 allele on linkage group (LG) G and the Rhg4 allele on LG A2 are important in conditioning resistance. Markers closely linked to the Rhg4 locus were used previously to screen a library of bacterial artificial chromosome (BAC) clones from susceptible 'Williams 82' and identified a single 150-kb BAC, Gm_ISb001_056_G02 (56G2). End-sequenced subclones positioned onto a restriction map provided landmarks for identifying the corresponding region from a BAC library from accession PI 437654 with broad resistance to SCN. Seventy-three PI 437654 BACs were assigned to contigs based upon HindIII restriction fragment profiles. Four contigs represented the PI 437654 counterpart of the 'Williams 82' BAC, with PCR assays connecting these contigs. Some of the markers on the PI 437654 contigs are separated by a greater physical distance than in the 'Williams 82' BAC and some primers amplify bands from BACs in the mid-portion of the connected PI 437654 BAC contigs that are not amplified from the 'Williams 82' BAC. These observations suggest that there is an insertion in the PI 437654 genome relative to the 'Williams 82' genome in the Rhg4 region.Key words: BAC, deletion, insertion, resistance gene, soybean cyst nematode.

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Development of a sequence-based reference physical map of pea (Pisum sativum L.)

10.1101/518563 ◽

2019 ◽

Author(s):

Krishna Kishore Gali ◽

Bunyamin Tar’an ◽

Mohammed-Amin Madoui ◽

Edwin van der Vossen ◽

Jan van Oeveren ◽

...

Keyword(s):

Genome Sequence ◽

Positional Cloning ◽

Physical Map ◽

Repetitive Sequences ◽

Bac Library ◽

Artificial Chromosome ◽

Bac Clones ◽

Pooled Dna ◽

Mapping Technology ◽

Generation Sequencing

AbstractWhole genome profiling (WGP) is a sequence-based physical mapping technology and uses sequence tags generated by next generation sequencing for construction of bacterial artificial chromosome (BAC) contigs of complex genomes. The physical map provides a framework for assembly of genome sequence and information for localization of genes that are difficult to find through positional cloning. To address the challenges of accurate assembly of the pea genome (~4.2 GB of which approximately 85% is repetitive sequences), we have adopted the WGP technology for assembly of a pea BAC library. Multi-dimensional pooling of 295,680 BAC clones and sequencing the ends of restriction fragments of pooled DNA generated 1,814 million high quality reads, of which 825 million were deconvolutable to 1.11 million unique WGP sequence tags. These WGP tags were used to assemble 220,013 BACs into contigs. Assembly of the BAC clones using the modified Fingerprinted Contigs (FPC) program has resulted in 13,040 contigs, consisting of 213,719 BACs, and 6,294 singleton BACs. The average contig size is 0.33 Mbp and the N50 contig size is 0.62 Mbp. WGPTM technology has proved to provide a robust physical map of the pea genome, which would have been difficult to assemble using traditional restriction digestion based methods. This sequence-based physical map will be useful to assemble the genome sequence of pea. Additionally, the 1.1 million WGP tags will support efficient assignment of sequence scaffolds to the BAC clones, and thus an efficient sequencing of BAC pools with targeted genome regions of interest.

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DECONVOLUTING BAC–GENE RELATIONSHIPS USING A PHYSICAL MAP

Journal of Bioinformatics and Computational Biology ◽

10.1142/s0219720008003564 ◽

2008 ◽

Vol 06 (03) ◽

pp. 603-622

Author(s):

YONGHUI WU ◽

LAN LIU ◽

TIMOTHY J. CLOSE ◽

STEFANO LONARDI

Keyword(s):

Physical Map ◽

Bac Library ◽

Artificial Chromosome ◽

Hybridization Experiment ◽

Pooling Design ◽

Bac Clones ◽

Hybridization Data ◽

A Genome ◽

The Right ◽

Combinatorial Pooling

Deconvolution of relationships between bacterial artificial chromosome (BAC) clones and genes is a crucial step in the selective sequencing of regions of interest in a genome. It often includes combinatorial pooling of unique probes obtained from the genes (unigenes), and screening of the BAC library using the pools in a hybridization experiment. Since several probes can hybridize to the same BAC, in order for the deconvolution to be achievable the pooling design has to be able to handle a large number of positives. As a consequence, smaller pools need to be designed, which in turn increases the number of hybridization experiments, possibly making the entire protocol unfeasible. We propose a new algorithm that is capable of producing high-accuracy deconvolution even in the presence of a weak pooling design, i.e. when pools are rather large. The algorithm compensates for the decrease of information in the hybridization data by taking advantage of a physical map of the BAC clones. We show that the right combination of combinatorial pooling and our algorithm not only dramatically reduces the number of pools required, but also successfully deconvolutes the BAC–gene relationships with almost perfect accuracy. Software is available on request from the first author.

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A bacterial artificial chromosome library for ‘Jefferson’ hazelnut and identification of clones associated with eastern filbert blight resistance and pollen–stigma incompatibility

Genome ◽

10.1139/g11-048 ◽

2011 ◽

Vol 54 (10) ◽

pp. 862-867 ◽

Cited By ~ 5

Author(s):

Vidyasagar R. Sathuvalli ◽

Shawn A. Mehlenbacher

Keyword(s):

Bacterial Artificial Chromosome ◽

Random Amplified Polymorphic Dna ◽

Bac Library ◽

Artificial Chromosome ◽

Physical Distance ◽

Resistance Locus ◽

S Locus ◽

Insert Size ◽

Eastern Filbert Blight ◽

Corylus Avellana L

European hazelnut ( Corylus avellana L.) is the only economically important nut crop in the family Betulaceae. Because of its small genome size (~385 Mb / 1C), relatively short life cycle, availability of a dense linkage map, and amenability to transformation by Agrobacterium, the European hazelnut could serve as a model plant for the Betulaceae. Here we report the construction of a bacterial artificial chromosome (BAC) library for ‘Jefferson’ hazelnut using the cloning enzyme MboI and the vector pECBAC1 (BamHI site). The library consists of 39 936 clones arrayed in 104 384-well microtitre plates with a mean insert size of 117 kb. The genomic coverage of the library is estimated to be about 12 genome equivalents. This library provides a valuable resource for the map-based cloning of two important genes, the resistance gene from ‘Gasaway’ that confers resistance to eastern filbert blight caused by the fungus Anisogramma anomala (Peck) E. Müller and the S locus that controls pollen–stigma incompatibility. Fine-resolution mapping near the two loci was carried out using random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) markers. Fine mapping at the disease resistance locus showed that markers W07-375 and X01-825 flanked the resistance locus at distances of 0.06 and 0.05 cM, respectively. The S locus is flanked by markers 204-950 and KG819-200 at distances of 0.14 and 0.24 cM, respectively. Assuming that 1 cM corresponds to a physical distance of 430 kb, it will take approximately two to three chromosome walks to assemble BAC contigs that span both loci.

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Construction of a bacterial artificial chromosome (BAC) library for potato molecular cytogenetics research

Genome ◽

10.1139/g99-099 ◽

2000 ◽

Vol 43 (1) ◽

pp. 199-204 ◽

Cited By ~ 32

Author(s):

Junqi Song ◽

Fenggao Dong ◽

Jiming Jiang

Keyword(s):

Bacterial Artificial Chromosome ◽

Physical Map ◽

Molecular Cytogenetics ◽

Bac Library ◽

Artificial Chromosome ◽

Average Insert Size ◽

Chromosome Identification ◽

Insert Size ◽

Potato Genome ◽

Bac Clones

Lack of reliable techniques for chromosome identification is the major obstacle for cytogenetics research in plant species with large numbers of small chromosomes. To promote molecular cytogenetics research of potato (Solanum tuberosum, 2n = 4x = 48) we developed a bacterial artificial chromosome (BAC) library of a diploid potato species S. bulbocastanum. The library consists of 23 808 clones with an average insert size of 155 kb, and represents approximately 3.7 equivalents to the potato genome. The majority of the clones in the BAC library generated distinct signals on specific potato chromosomes using fluorescence in situ hybridization (FISH). The hybridization signals provide excellent cytological markers to tag individual potato chromosomes. We also demonstrated that the BAC clones can be mapped to specific positions on meiotic pachytene chromosomes. The excellent resolution of pachytene FISH can be used to construct a physical map of potato by mapping molecular marker-targeted BAC clones on pachytene chromosomes. Key words: potato, BAC library, chromosome identification, physical mapping, molecular cytogenetics.

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Genomic organization of the 260 kb surrounding the waxy locus in a Japonica rice

Genome ◽

10.1139/g99-068 ◽

1999 ◽

Vol 42 (6) ◽

pp. 1121-1126 ◽

Cited By ~ 4

Author(s):

Hironori Nagano ◽

Lihua Wu ◽

Shinji Kawasaki ◽

Yuji Kishima ◽

Yoshio Sano

Keyword(s):

Genomic Organization ◽

Physical Map ◽

Repetitive Sequences ◽

Bac Library ◽

Artificial Chromosome ◽

Molecular Organization ◽

Japonica Rice ◽

Class 1 ◽

Average Size ◽

Waxy Locus

The present study was carried out to characterize the molecular organization in the vicinity of the waxy locus in rice. To determine the structural organization of the region surrounding waxy, contiguous clones covering a total of 260 kb were constructed using a bacterial artificial chromosome (BAC) library from the Shimokita variety of Japonica rice. This map also contains 200 overlapping subclones, which allowed construction of a fine physical map with a total of 64 HindIII sites. During the course of constructing the map, we noticed the presence of some repeated regions which might be related to transposable elements. We divided the 260-kb region into 60 segments (average size of 5.7 kb) to use as probes to determine their genomic organization. Hybridization patterns obtained by probing with these segments were classified into four types: class 1, a single or a few bands without a smeared background; class 2, a single or a few bands with a smeared background; class 3, multiple discrete bands without a smeared background; and class 4, only a smeared background. These classes constituted 6.5%, 20.9%, 3.7%, and 68.9% of the 260-kb region, respectively. The distribution of each class revealed that repetitive sequences are a major component in this region, as expected, and that unique sequence regions were mostly no longer than 6 kb due to interruption by repetitive sequences. We discuss how the map constructed here might be a powerful tool for characterization and comparison of the genome structures and the genes around the waxy locus in the Oryza species.Key words: BAC library, genomic organization, physical map, rice (Oryza sativa), the waxy locus.

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A bacterial artificial chromosome (BAC) library of sugar beet and a physical map of the region encompassing the bolting gene B

Molecular Genetics and Genomics ◽

10.1007/s00438-003-0821-7 ◽

2003 ◽

Vol 269 (1) ◽

pp. 126-136 ◽

Cited By ~ 19

Author(s):

U. Hohmann ◽

G. Jacobs ◽

A. Telgmann ◽

R. M. Gaafar ◽

S. Alam ◽

...

Keyword(s):

Bacterial Artificial Chromosome ◽

Sugar Beet ◽

Physical Map ◽

Bac Library ◽

Artificial Chromosome

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Construction of a long-range YAC physical map spanning the 10-cM region between the markers D18Mit109 and D18Mit68 on mouse proximal chromosome 18

Genome ◽

10.1139/g99-124 ◽

2000 ◽

Vol 43 (3) ◽

pp. 427-433

Author(s):

Ssucheng J Hsu ◽

Robert P Erickson

Keyword(s):

Yeast Artificial Chromosome ◽

Physical Map ◽

Artificial Chromosome ◽

Physical Distance ◽

Chromosome 18 ◽

Sequence Tagged Sites ◽

Npc1 Gene ◽

Niemann Pick ◽

Yac Contigs ◽

Physical Order

Four yeast artificial chromosome (YAC) contigs, physically~8 Mb, have been constructed spanning a 10-cM region on mouse proximal chromosome 18 and include the sites of 21 known genes, including those near the twirler (Tw) locus and the recently isolated Niemann-Pick type C1 (npc1) gene, formerly designated as the spm locus. This physical map consists of 49 YAC clones that cover roughly 15% of the chromosome. The physical order of 38 microsatellite sequence-tagged sites (STSs) could be assembled and confirmed based on their presence or absence in individual YACs, from proximal D18Mit109 through distal D18Mit68. These YACs provide an important resource for the further characterization and identification of known and unknown genes. The physical map has been integrated with our previously published genetic linkage map and showed an average genetic to physical distance of cM/Mb > 1.1.Key words: Mus musculus, chromosome 18, YAC contigs, physical mapping, Niemann-Pick type C1.

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484 Constructing the BAC Contig for the Evergreen Gene Region in Peach [Prunus persica (L.) Batsch]

HortScience ◽

10.21273/hortsci.35.3.477e ◽

2000 ◽

Vol 35 (3) ◽

pp. 477E-478

Author(s):

Ying Wang ◽

Laura L. George ◽

Gregory L. Reighard ◽

Ralph Scorza ◽

Albert G. Abbott

Keyword(s):

Prunus Persica ◽

Physical Map ◽

Single Gene ◽

Bac Library ◽

Gene Region ◽

Average Insert Size ◽

Insert Size ◽

Bac Clone ◽

Bac Contig ◽

Peach Genome

Evergreen genotypes of peach [Prunus persica (L.) Batsch] have been identified in Mexico, where terminal growth on evergreen trees is continuous under favorable environmental conditions. This evergreen trait in peach is controlled by one single gene (evg), and this evergreen condition is homozygous recessive. Four dominant AFLP markers, EAT/MCAC, ETT/MCCA2, EAT/MCTA, and ETT/MACC, were found to be tightly linked to the evg locus at 1 cM, 4.6 cM, 5.8 cM, and 11 cM, respectively. All four markers were sequenced and identified. A peach BAC library was constructed by using the pBeloBAC11 vector for building the physical map for the evg gene. This library represents four times the coverage of the peach genome with the average insert size of 50 to 70 kb. The EAT/MCAC AFLP marker fragment was used for screening the peach BAC library. A single BAC clone, 18F12, was confirmed to contain this fragment. The final BAC contig for this evg gene region and the potential homology between peach and Arabidopsis thaliana will be presented and discussed.

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