Generation of sequence-tagged sites from Xp22.3 by isolating commonAlu-PCR products of radiation hybrids retaining overlapping human X chromosome fragments

Abstract A physical map of the euchromatic X chromosome of Drosophila melanogaster has been constructed by assembling contiguous arrays of cosmids that were selected by screening a library with DNA isolated from microamplified chromosomal divisions. This map, consisting of 893 cosmids, covers approximately 64% of the euchromatic part of the chromosome. In addition, 568 sequence tagged sites (STS), in aggregate representing 120 kb of sequenced DNA, were derived from selected cosmids. Most of these STSs, spaced at an average distance of approximately 35 kb along the euchromatic region of the chromosome, represent DNA tags that can be used as entry points to the fruitfly genome. Furthermore, 42 genes have been placed on the physical map, either through the hybridization of specific probes to the cosmids or through the fact that they were represented among the STSs. These provide a link between the physical and the genetic maps of D. melanogaster. Nine novel genes have been tentatively identified in Drosophila on the basis of matches between STS sequences and sequences from other species.

Download Full-text

Clonal origin of non-medullary thyroid tumours assessed by non-random X-chromosome inactivation

Acta Endocrinologica ◽

10.1530/eje.0.1460027 ◽

2002 ◽

pp. 27-33 ◽

Cited By ~ 31

Author(s):

S Moniz ◽

AL Catarino ◽

AR Marques ◽

B Cavaco ◽

L Sobrinho ◽

...

Keyword(s):

Thyroid Carcinoma ◽

X Chromosome ◽

X Chromosome Inactivation ◽

Chromosome Inactivation ◽

Cag Repeats ◽

Surrounding Tissue ◽

Normal Thyroid ◽

Medullary Thyroid ◽

Pcr Products ◽

Clonal Origin

OBJECTIVE: X-chromosome inactivation analysis was performed in order to assess the clonal origin of non-medullary thyroid tumours and to distinguish between multicentricity and multifocality in multiple papillary thyroid carcinoma (PTC). METHODS: One hundred and thirteen tumour samples from 31 patients with isolated PTC, 16 patients with multinodular PTC, 14 patients with follicular thyroid adenoma (FTA) and 15 patients with follicular thyroid carcinoma (FTC) were collected. The corresponding normal thyroid tissues were analysed, and in 14 cases, tumour-surrounding tissue was also studied. Genomic DNA was digested with HpaII and HhaI previous to PCR amplification of the polymorphic CAG repeat, on exon 1 of the human androgen receptor gene (HUMARA). PCR products were analysed by denaturing gel electrophoresis, silver staining and densitometric analysis. PCR products were also used to determine the number of CAG repeats of patients with isolated PTC, FTA, FTC and of 41 healthy volunteers. RESULTS: Heterozygosity for the HUMARA polymorphism was found in 64/76 (84%) cases. Lyonization of the thyroid was observed in 15/76 (20%) cases, which were excluded from clonal analysis. Except for two cases of isolated PTC, all tumour samples studied presented monoclonal X-inactivation patterns, while normal thyroid tissue was polyclonal. Monoclonal patterns were also found in 4/14 tumour-surrounding tissues. No difference was found in the length of CAG alleles between patients and controls. Of eight informative cases of multinodular PTC, three showed evidence of multicentricity and five revealed patterns consistent with multifocality. CONCLUSIONS: Both isolated and multinodular PTC as well as FTA and FTC are of monoclonal origin. Our results also suggest that approximately one-third of multiple PTC have an independent origin for the different nodules (multicentricity). Monoclonality was also found in tissues surrounding some PTC nodules. No association was found between the length of CAG alleles and thyroid malignancies.

Download Full-text

DOP–PCR based painting of rye chromosomes in a wheat background

Genome ◽

10.1139/gen-2014-0110 ◽

2014 ◽

Vol 57 (9) ◽

pp. 473-479 ◽

Cited By ~ 6

Author(s):

Chuanliang Deng ◽

Lili Bai ◽

Shufen Li ◽

Yingxin Zhang ◽

Xiang Li ◽

...

Keyword(s):

Dna Sequences ◽

Chromosome Painting ◽

Pcr Amplification ◽

Translocation Line ◽

Addition Line ◽

Metaphase Chromosomes ◽

Application Range ◽

Degenerate Oligonucleotide ◽

Pcr Products ◽

Chromosome Fragments

To determine the appropriateness of chromosome painting for identifying genomic elements in rye, we microdissected the 1R and 1RS chromosomes from rye (Secale cereale L. var. King II) and wheat–rye addition line 1RS, respectively. Degenerate oligonucleotide primed – polymerase chain reaction (DOP–PCR) amplification of 1R and 1RS products from dissected chromosomes were used as probes to hybridize to metaphase chromosomes of rye, wheat–rye addition lines 1R and 1RS, translocation line 1RS.1BL, and allohexaploid triticale. The results showed that (i) the hybridization signal distribution patterns on rye chromosomes using 1R-derived DOP–PCR products as the probe were similar to those using 1RS-derived DOP–PCR products as the probe; (ii) 1R and (or) 1RS could not be distinguished from other rye chromosomes solely by the hybridization patterns using 1R- and (or) 1RS-derived DOP–PCR products as the probe; (iii) rye chromosomes and (or) rye chromosome fragments could be clearly identified in wheat–rye hybrids using either 1R- or 1RS-derived DOP–PCR products as the probe and could be more accurate in the nontelomeric region than using genomic in situ hybridization (GISH). Our results suggested that 1R- and (or) 1RS-derived DOP–PCR products contain many repetitive DNA sequences, are similar on different rye chromosomes, are R-genome specific, and can be used to identify rye chromosomes and chromosome fragments in wheat–rye hybrids. Our research widens the application range of chromosome painting in plants.

Download Full-text

Number and size of human X chromosome fragments transferred to mouse cells by chromosome-mediated gene transfer.

Molecular and Cellular Biology ◽

10.1128/mcb.1.5.439 ◽

1981 ◽

Vol 1 (5) ◽

pp. 439-448 ◽

Cited By ~ 12

Author(s):

A S Olsen ◽

O W McBride ◽

D E Moore

Keyword(s):

Gene Transfer ◽

X Chromosome ◽

Deoxyribonucleic Acid ◽

Single Copy ◽

Mouse Cell ◽

Unique Sequence ◽

Chromosome Fragments ◽

Mouse Cells ◽

Chromosomal Sequences ◽

Kinetics Of

Labeled probes of unique-sequence human X chromosomal deoxyribonucleic acid, prepared by two different procedures, were used to measure the amount of human X chromosomal deoxyribonucleic acid in 12 mouse cell lines expressing human hypoxanthine phosphoribosyltransferase after chromosome-mediated gene transfer. The amount of X chromosomal deoxyribonucleic acid detected by this procedure ranged from undetectable levels in the three stable transformants and some unstable transformants examined to about 20% of the human X chromosome in two unstable transformants. Reassociation kinetics of the X chromosomal probe with deoxyribonucleic acid from the two unstable transformants containing 15 to 20% of the human X chromosome indicate that a single copy of these sequences is present. In one of these lines, the X chromosomal sequences exist as multiple fragments which were not concordantly segregated when the cells were selected for loss of hprt.

Download Full-text

Occurrence and inheritance of microsatellites in Pinus radiata

Genome ◽

10.1139/g94-138 ◽

1994 ◽

Vol 37 (6) ◽

pp. 977-983 ◽

Cited By ~ 78

Author(s):

David N. Smith ◽

Michael E. Devey

Keyword(s):

Pinus Radiata ◽

Population Genetic ◽

Dna Marker ◽

Genomic Library ◽

Mendelian Inheritance ◽

Dinucleotide Repeats ◽

Sequence Tagged Sites ◽

Pcr Products ◽

Conifer Genome ◽

Highly Repetitive Dna

Microsatellites are an important class of DNA marker because of their abundance and length hypervariability. As part of a project mapping the Pinus radiata genome, we have characterized some of the microsatellites in this species. Southern blots were screened with oligonucleotide probes [(CA)10, (GA)10, (GAA)9, (CAA)8, (CAC)5, (GACA)4] to assess their abundance. CA and GA were the most abundant microsatellites, while GAA was least abundant. A genomic library in lambda ZAP, covering 9 × 104 kb, was screened with a combined poly(CA) + poly(GA) probe and yielded 120 positives, approximately one CA or GA microsatellite every 750 kb of the P. radiata genome. It was found that 25% of the positives were embedded within highly repetitive DNA. Four of the five subclones sequenced contained compound microsatellites, with TA predominating as the additional repeat. Segregation analysis of PCR products for two microsatellites, PR4.6 and PR9.3, in 96 progeny of a controlled out-cross verified simple Mendelian inheritance. Both loci are highly polymorphic with Polymorphism Information Content values of 0.63 and 0.70 for PR4.6 and PR9.3, respectively. These results indicate that microsatellites are abundant in a conifer genome and can be valuable markers for pine mapping, fingerprinting, and population genetic studies.Key words: microsatellites, dinucleotide repeats, conifer genome, sequence tagged sites.

Download Full-text

Number and size of human X chromosome fragments transferred to mouse cells by chromosome-mediated gene transfer

Molecular and Cellular Biology ◽

10.1128/mcb.1.5.439-448.1981 ◽

1981 ◽

Vol 1 (5) ◽

pp. 439-448

Author(s):

A S Olsen ◽

O W McBride ◽

D E Moore

Keyword(s):

Gene Transfer ◽

X Chromosome ◽

Deoxyribonucleic Acid ◽

Single Copy ◽

Mouse Cell ◽

Unique Sequence ◽

Chromosome Fragments ◽

Mouse Cells ◽

Chromosomal Sequences ◽

Kinetics Of

Labeled probes of unique-sequence human X chromosomal deoxyribonucleic acid, prepared by two different procedures, were used to measure the amount of human X chromosomal deoxyribonucleic acid in 12 mouse cell lines expressing human hypoxanthine phosphoribosyltransferase after chromosome-mediated gene transfer. The amount of X chromosomal deoxyribonucleic acid detected by this procedure ranged from undetectable levels in the three stable transformants and some unstable transformants examined to about 20% of the human X chromosome in two unstable transformants. Reassociation kinetics of the X chromosomal probe with deoxyribonucleic acid from the two unstable transformants containing 15 to 20% of the human X chromosome indicate that a single copy of these sequences is present. In one of these lines, the X chromosomal sequences exist as multiple fragments which were not concordantly segregated when the cells were selected for loss of hprt.

Download Full-text

Establishment of Sequence-Tagged Sites on 15q11-q13 byAlu-Vector PCR Cloning of Yac-Generated Fragments

Disease Markers ◽

10.1155/1996/167830 ◽

1996 ◽

Vol 12 (4) ◽

pp. 241-246

Author(s):

W. S. Kim ◽

Z. M. Deng ◽

N. T. Nassif ◽

A. Smith ◽

R. J. Trent

Keyword(s):

Human Chromosome ◽

Angelman Syndrome ◽

Chromosome 15 ◽

Loss Of Function ◽

Pcr Cloning ◽

Artificial Chromosomes ◽

Sequence Tagged Sites ◽

Pcr Products ◽

Yeast Artificial Chromosomes

Angelman syndrome (AS) is caused by the loss of function of undefined gene(s) on human chromosome 15. The majority of subjects have deletions involving maternally-derived chromosome 15q II-q 13, and the shortest region of deletion overlap (SRO) has been localized to the region between D15S10 and D15S113. In this study, yeast artificial chromosomes (YACs), 6G-D4, 9H-D2 and 37D-F9, mapping within the AS SRO, were isolated from the ICI Y AC library.Alu-vector PCR products were amplified from the YACs and from YACs A229A2 and A33FI 0 which had been obtained from the St. Louis Y AC library. The PCR products were cloned and sequenced, and three new sequence-tagged sites were generated within the AS SRO, facilitating the characterization of gene(s) involved in the Angelman syndrome.

Download Full-text