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1.
Poult Sci ; 81(11): 1644-6, 2002 Nov.
Article in English | MEDLINE | ID: mdl-12455590

ABSTRACT

Chromosome-specific painting probes and libraries were developed for chicken Macrochromosomes 1, 2, 3, and 4 by chromosome microisolation and microcloning. Fluorescent in situ hybridization results using the painting probes on normal chicken metaphase chromosomes indicated the purity and specificity of each probe. Chromosome-specific libraries for chicken Macrochromosomes 1, 2, 3, and 4 were prepared in a phage vector. Fifty-two additional unique microsatellite markers of the (AC)n type were developed from these chromosome-specific libraries. These markers were mapped on the East Lansing reference population to increase the marker density on the four macrochromosomes. Results of the current study suggest that development of markers from chromosome-specific libraries is very useful for constructing high-density linkage maps for chicken macrochromosomes.


Subject(s)
Chickens/genetics , Chromosome Mapping/veterinary , Microsatellite Repeats , Animals , Cloning, Molecular , DNA Primers/chemistry , DNA Probes/chemistry , Gene Library , Genetic Linkage , In Situ Hybridization, Fluorescence/veterinary , Metaphase/genetics
2.
Anim Genet ; 32(3): 152-5, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11493264

ABSTRACT

A chromosome-specific library was developed for Bos taurus autosome 11 by chromosome microdissection and microcloning using a bovine primary fibroblast culture, obtained from a t(X;23) heifer, that spontaneously developed a translocation chromosome involving bovine chromosome 11. The library was screened using (AC)12 oligos, positive clones selected, sequenced and primers developed to generate bovine chromosome 11-specific microsatellite markers. This study suggests that chromosome-specific libraries have great potential for development of microsatellite markers for the construction of marker-saturated linkage maps for each chromosome.


Subject(s)
Cattle/genetics , Chromosome Mapping , Microsatellite Repeats , Translocation, Genetic , X Chromosome , Animals , Female , Gene Library , Genetic Markers
3.
Oncogene ; 20(21): 2671-82, 2001 May 10.
Article in English | MEDLINE | ID: mdl-11420679

ABSTRACT

A number of non-virally and non-chemically immortalized chicken embryo fibroblast (CEF) cells have been established recently in continuous cell culture. All immortal CEF cells tested showed common genetic alterations in the expression patterns of p53 and E2F-1 mRNA and protein which were down- and up-regulated, respectively. The biological effects of differentially regulated p53 and E2F-1 were determined by reporter gene transcriptional activity assays, DNA binding assays, and Northern blot analysis of the expression patterns of down-stream genes. In addition, expression of most of the cyclin genes was up-regulated in immortal CEF cells, which may be associated with the rapid cell division rates and serum-independent growth patterns seen in immortal CEF cells. The telomeric lengths and chromosome integrity were maintained in all immortal CEF cell lines without detectable telomerase activity. Although the functional inactivations of the p53 and Rb regulatory pathways are known to be common events for cellular immortalization, the genetic changes leading to alteration of p53 and E2F-1 function through transcriptional and post-transcriptional regulation seem to be unique in immortal CEF cells.


Subject(s)
Carrier Proteins , Cell Cycle Proteins , DNA-Binding Proteins , Transcription Factors/physiology , Tumor Suppressor Protein p53/physiology , Cell Division/physiology , Cell Line, Transformed , Chromosomes/physiology , Cyclins/biosynthesis , Cyclins/genetics , E2F Transcription Factors , E2F1 Transcription Factor , Fibroblasts/metabolism , Fibroblasts/physiology , Gene Silencing , Genes, Reporter/genetics , Genes, cdc/physiology , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Retinoblastoma-Binding Protein 1 , Telomere/physiology , Transcription Factors/biosynthesis , Transcription Factors/genetics , Transcription, Genetic , Tumor Suppressor Protein p53/biosynthesis , Tumor Suppressor Protein p53/genetics
4.
Mol Reprod Dev ; 59(1): 67-77, 2001 May.
Article in English | MEDLINE | ID: mdl-11335948

ABSTRACT

Testicular activity and semen characteristics of bulls carrying an X-autosome translocation t(Xp +;23q-) revealed all stages of spermatogenesis although their semen consisted of few and, exclusively, of malformed spermatozoa. Chromosome painting on metaphase spreads of their mother and synaptonemal complex analysis on these and normal bulls were carried out to test whether the location and meiotic pairing behaviour of the rearranged segments could have contributed to the sperm head malformation and oligospermia in our X-autosome translocation (X-AT) carrier bulls. Spermatocytes of X-AT carriers displayed the rearranged chromosomes in a univalent-trivalent association, with 23q- always remaining as a univalent and Xp + in synapsis with normal chromosome 23 and the Y chromosome. Chromosome painting studies to test whether the total absence of meiocytes showing a quadrivalent is due to the non-reciprocal nature of this translocation, identified Xp sequence homology with the distal end of 23q- confirming its relocation to the terminal segment of 23q-. Our synaptonemal complex analyses also confirmed that the bovine pseudo-autosomal region (PAR) is at the distal ends of Xq and Yp and further revealed that over 85% of spermatocytes of X-AT carriers (and up to 13% of spermatocytes of normal bulls) sustain a Y-axis break adjacent to the PAR. Although the exact cause of a Y-axis break in bovine spermatocytes is not known at present, we believe that the break and possible loss of Yq in such high proportions of spermatocytes of X-AT carriers could have contributed to the sperm head malformation and oligospermia in our X-AT carrier bulls.


Subject(s)
Cattle/genetics , Sperm Head/ultrastructure , Spermatozoa/cytology , Synaptonemal Complex/ultrastructure , Translocation, Genetic , X Chromosome/genetics , Animals , Cattle/physiology , Cells, Cultured , Chromosome Painting , Chromosomes/genetics , Chromosomes/ultrastructure , Female , Fibroblasts , Humans , Male , Semen/cytology , Sperm Count , Spermatogenesis , Testis/cytology , X Chromosome/ultrastructure
5.
Am J Pathol ; 156(3): 1099-107, 2000 Mar.
Article in English | MEDLINE | ID: mdl-10702426

ABSTRACT

The Smyth line (SL) chicken, an animal model for autoimmune human vitiligo, is characterized by a spontaneous posthatch pigment loss, determined to be the result of an autoimmune phenomenon. Because endogenous virus (EV) genes have been reported to be associated with a number of autoimmune diseases of human and animal models, we designed this experiment to investigate the role of EV in the SL vitiligo by using the complete sequence of Rous-associated virus-2 as a probe for EV. An F(2) resource population was developed by the matings of SL and parental control (BL) chickens. Linkage disequilibrium between vitiligo and EV was apparent (16.2-kb SacI fragment, P

Subject(s)
Autoimmune Diseases/virology , Chickens/virology , Disease Models, Animal , Endogenous Retroviruses/genetics , Genes, Viral , Vitiligo/virology , Animals , Autoimmune Diseases/genetics , Azacitidine/pharmacology , Blotting, Northern , Chickens/genetics , DNA/analysis , DNA/chemistry , DNA Fingerprinting , DNA Methylation , Deoxyribonuclease HindIII/metabolism , Deoxyribonucleases, Type II Site-Specific/metabolism , Female , Fibroblasts/drug effects , Fibroblasts/metabolism , In Situ Hybridization, Fluorescence , Male , Pedigree , Polymorphism, Restriction Fragment Length , RNA Viruses , RNA, Viral/analysis , Vitiligo/genetics
6.
Anim Genet ; 30(4): 251-5, 1999 Aug.
Article in English | MEDLINE | ID: mdl-10467699

ABSTRACT

To develop additional microsatellite (MS) markers in the region of the porcine skeletal muscle ryanodine receptor gene (RYR1), a microdissected genomic library was generated from the proximal half of the q arm of swine chromosome 6. Purified DNA was restriction enzyme-digested, ligated to oligonucleotide adaptors and amplified by PCR using primers complementary to the adaptor sequences. The purity of the amplified products and boundaries of the microdissected chromosomal region were verified by fluorescence in situ hybridization. (CA)n-containing sequences were then identified in a small insert genomic library generated from the PCR-amplified microdissected DNA. Oligonucleotide primers were developed for the PCR amplification of 30 of the 46 (CA)n repeat-containing clones, which were subsequently used to amplify DNA isolated from unrelated pigs of different breeds to determine the informativeness of these MS markers. Twenty-two of these MS markers were genotyped on the University of Illinois Yorkshire x Meishan swine reference population. These 22 markers were all assigned within a 50.7-CM region of the swine chromosome 6 linkage map, indicating the specificity of the microdissected library.


Subject(s)
Genomic Library , Microsatellite Repeats , Swine/genetics , Animals , Base Sequence , Chromosome Mapping , Chromosomes/genetics , DNA Primers/genetics , Genetic Linkage , Genotype , Polymerase Chain Reaction , Polymorphism, Genetic , Ryanodine Receptor Calcium Release Channel/genetics
7.
J Hered ; 88(3): 247-9, 1997.
Article in English | MEDLINE | ID: mdl-9183849

ABSTRACT

We have developed a chicken (Gallus domesticus) Z-chromosome-specific DNA library in a phage vector by means of chromosome microisolation and microcloning. The chromosomal origin, specificity, and purity was evaluated by fluorescent in situ hybridization (FISH) on chicken metaphases. Heterologous chromosome painting using this Z-chromosome-specific probe on turkey (Meleagris gallopavo) metaphases identified its homologous Z-chromosome, under the same stringent conditions as that used in the chicken, indicating a high degree of Z-chromosome sequence homology among these two species. This chicken Z-chromosome library will facilitate the development of Z-chromosome-specific DNA markers that will be useful for genetic mapping in the domestic chicken and related avian species. The Z-chromosome-specific DNA probe will also be useful for studies pertaining to the sex chromosome evolution in avian species.


Subject(s)
Chromosomes , DNA , Gene Library , Animals , Chickens/genetics , Metaphase , Turkey
8.
Mamm Genome ; 8(1): 16-20, 1997 Jan.
Article in English | MEDLINE | ID: mdl-9021142

ABSTRACT

Genotypic data for 56 microsatellites (ms) generated from maternal full sib families nested within paternal half sib pedigrees were used to construct a linkage map of the bovine X Chromosome (Chr) (BTX) that spans 150 cM (ave. interval 2.7 cM). The linkage map contains 36 previously unlinked ms; seven generated from a BTXp library. Genotypic data from these 36 ms was merged into an existing linkage map to more than double the number of informative BTX markers. A male specific linkage map of the pseudoautosomal region was also constructed from five ms at the distal end of BTXq. Four informative probes physically assigned by fluorescence in situ hybridization defined the extent of coverage, confirmed the position of the pseudoautosomal region on the q-arm, and identified a 4.1-cM marker interval containing the centromere of BTX.


Subject(s)
Cattle/genetics , Chromosome Mapping/veterinary , X Chromosome , Animals , Female , Genetic Linkage , Male , Microsatellite Repeats
9.
Cytogenet Cell Genet ; 76(1-2): 27-33, 1997.
Article in English | MEDLINE | ID: mdl-9154119

ABSTRACT

To facilitate the identification of microsatellite genetic markers from a single swine chromosome, chromosome microisolation and microcloning have been used to generate a swine chromosome 6-specific DNA library. Ten copies of swine chromosome 6 were scraped from metaphase spreads, ligated to custom-prepared adaptors, and amplified by PCR. The purity of the amplified product was verified by fluorescent in situ hybridization. The utility of the chromosome painting probe for heterologous painting was demonstrated and confirmed that swine chromosome 6 is syntenic to human chromosomes 1p and 19q. A small insert genomic library of 1.39 x 10(6) clones was generated from the PCR-amplified chromosome 6 genomic DNA and screened for (GT)n microsatellite genetic markers. Nine (GT)n microsatellite markers were developed and genotyped on a Yorkshire x Meishan swine reference family. All nine markers genetically mapped to chromosome 6, confirming the purity of the microisolation method. The method used here should be adaptable to the microdissection of subchromosomal regions of not only the swine genome but also other livestock genomes.


Subject(s)
Gene Library , Swine/genetics , Animals , Humans , Microsatellite Repeats
10.
Proc Natl Acad Sci U S A ; 93(8): 3450-4, 1996 Apr 16.
Article in English | MEDLINE | ID: mdl-8622956

ABSTRACT

The X chromosome linkage group is conserved in placental mammals. However, X chromosome morphological differences, due to internal chromosome rearrangements, exist among mammalian species. We have developed bovine chromosome painting probes for Xp and Xq to assess segment homologies between the submetacentric bovine X chromosome and the acrocentric sheep and goat X chromosomes. These painting probes and their corresponding DNA libraries were developed by chromosome micromanipulation, DNA micropurification, microcloning, and PCR amplification. The bovine Xp painting probe identified an interstitially located homologous segment in the sheep and goat Xq region, most probably resulting from chromosome inversion. Ten type II (microsatellite) markers obtained from the bovine Xq library and five other X chromosome assigned, but unlinked, markers were used to generate a linkage map for Xq spanning 89.4 centimorgans. The chromosome painting probes and molecular markers generated in this study would be useful for comparative mapping and tracing of internal X chromosome rearrangements in all ruminant species and would contribute to the understanding of mammalian sex chromosome evolution.


Subject(s)
Cattle/genetics , Genetic Linkage , Goats/genetics , Sheep/genetics , X Chromosome/genetics , Animals , Base Sequence , Chromosome Banding , Chromosome Mapping , DNA Primers/genetics , Female , Gene Library , Genetic Markers , In Situ Hybridization, Fluorescence , Molecular Probes , Molecular Sequence Data , Species Specificity , X Chromosome/ultrastructure
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