scholarly journals Engineering of Systematic Elimination of a Targeted Chromosome in Human Cells

2017 ◽  
Vol 2017 ◽  
pp. 1-5 ◽  
Author(s):  
Hiroshi Sato ◽  
Hiroki Kato ◽  
Haruyoshi Yamaza ◽  
Keiji Masuda ◽  
Huong Thi Nguyen Nguyen ◽  
...  
Keyword(s):  
Normal Cell ◽  
Expression Vector ◽  
Gene Dosage ◽  
Genetic Disorders ◽  
Human Cells ◽  
Chromosome 21 ◽  
Disease Phenotypes ◽  
Simplex Virus ◽  
Selection Of ◽  
Trisomic Cell

Embryonic trisomy leads to abortion or congenital genetic disorders in humans. The most common autosomal chromosome abnormalities are trisomy of chromosomes 13, 18, and 21. Although alteration of gene dosage is thought to contribute to disorders caused by extra copies of chromosomes, genes associated with specific disease phenotypes remain unclear. To generate a normal cell from a trisomic cell as a means of etiological analysis or candidate therapy for trisomy syndromes, we developed a system to eliminate a targeted chromosome from human cells. Chromosome 21 was targeted by integration of a DNA cassette in HeLa cells that harbored three copies of chromosome 21. The DNA cassette included two inverted loxP sites and a herpes simplex virus thymidine kinase (HSV-tk) gene. This system causes missegregation of chromosome 21 after expression of Cre recombinase and subsequently enables the selection of cells lacking the chromosome by culturing in a medium that includes ganciclovir (GCV). Cells harboring only two copies of chromosome 21 were efficiently induced by transfection of a Cre expression vector, indicating that this approach is useful for eliminating a targeted chromosome.

scholarly journals BK virus-plasmid expression vector that persists episomally in human cells and shuttles into Escherichia coli.

1984 ◽  
Vol 4 (8) ◽  
pp. 1551-1560 ◽  
Author(s):  
G Milanesi ◽  
G Barbanti-Brodano ◽  
M Negrini ◽  
D Lee ◽  
A Corallini ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Expression Vector ◽  
Selective Pressure ◽  
T Antigen ◽  
Bk Virus ◽  
Human Cells ◽  
Transformed Cells ◽  
Low Molecular Weight ◽  
Simplex Virus

We describe a novel expression vector, pBK TK-1, that persists episomally in human cells that can be shuttled into bacteria. This vector includes sequences from BK virus (BKV), the thymidine kinase (TK) gene of herpes simplex virus type 1, and plasmid pML-1. TK+-transformed HeLa and 143 B cells contained predominantly full-length episomes. There were typically 20 to 40 (HeLa) and 75 to 120 143 B vector copies per cell, although some 143 B transformants contained hundreds. Low-molecular-weight DNA from TK+-transformed cells introduced into Escherichia coli were recovered as plasmids that were indistinguishable from the input vector. Removal of selective pressure had no apparent effect upon the episomal status of pBK TK-1 molecules in TK+-transformed cells. BKV T antigen may play a role in episomal replication of pBK TK-1 since this viral protein was expressed in TK+ transformants and since a plasmid that contained only the BKV origin of replication was highly amplified in BKV-transformed human cells that synthesize BKV T antigen.

scholarly journals BK virus-plasmid expression vector that persists episomally in human cells and shuttles into Escherichia coli

1984 ◽  
Vol 4 (8) ◽  
pp. 1551-1560
Author(s):  
G Milanesi ◽  
G Barbanti-Brodano ◽  
M Negrini ◽  
D Lee ◽  
A Corallini ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Expression Vector ◽  
Selective Pressure ◽  
T Antigen ◽  
Bk Virus ◽  
Human Cells ◽  
Transformed Cells ◽  
Low Molecular Weight ◽  
Simplex Virus

We describe a novel expression vector, pBK TK-1, that persists episomally in human cells that can be shuttled into bacteria. This vector includes sequences from BK virus (BKV), the thymidine kinase (TK) gene of herpes simplex virus type 1, and plasmid pML-1. TK+-transformed HeLa and 143 B cells contained predominantly full-length episomes. There were typically 20 to 40 (HeLa) and 75 to 120 143 B vector copies per cell, although some 143 B transformants contained hundreds. Low-molecular-weight DNA from TK+-transformed cells introduced into Escherichia coli were recovered as plasmids that were indistinguishable from the input vector. Removal of selective pressure had no apparent effect upon the episomal status of pBK TK-1 molecules in TK+-transformed cells. BKV T antigen may play a role in episomal replication of pBK TK-1 since this viral protein was expressed in TK+ transformants and since a plasmid that contained only the BKV origin of replication was highly amplified in BKV-transformed human cells that synthesize BKV T antigen.

scholarly journals RTEL1 influences the abundance and localization of TERRA RNA

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Fiorella Ghisays ◽  
Aitor Garzia ◽  
Hexiao Wang ◽  
Claudia Canasto-Chibuque ◽  
Marcel Hohl ◽  
...  
Keyword(s):  
Cell Viability ◽  
Telomere Length ◽  
Human Cells ◽  
Helicase Domain ◽  
Telomere Repeat ◽  
Disease Phenotypes ◽  
G Quadruplex ◽  
Non Coding Rnas ◽  
Subtelomeric Regions

AbstractTelomere repeat containing RNAs (TERRAs) are a family of long non-coding RNAs transcribed from the subtelomeric regions of eukaryotic chromosomes. TERRA transcripts can form R-loops at chromosome ends; however the importance of these structures or the regulation of TERRA expression and retention in telomeric R-loops remain unclear. Here, we show that the RTEL1 (Regulator of Telomere Length 1) helicase influences the abundance and localization of TERRA in human cells. Depletion of RTEL1 leads to increased levels of TERRA RNA while reducing TERRA-containing R loops at telomeres. In vitro, RTEL1 shows a strong preference for binding G-quadruplex structures which form in TERRA. This binding is mediated by the C-terminal region of RTEL1, and is independent of the RTEL1 helicase domain. RTEL1 binding to TERRA appears to be essential for cell viability, underscoring the importance of this function. Degradation of TERRA-containing R-loops by overexpression of RNAse H1 partially recapitulates the increased TERRA levels and telomeric instability associated with RTEL1 deficiency. Collectively, these data suggest that regulation of TERRA is a key function of the RTEL1 helicase, and that loss of that function may contribute to the disease phenotypes of patients with RTEL1 mutations.

scholarly journals Genetics advances and learning disability

2000 ◽  
Vol 176 (1) ◽  
pp. 12-19 ◽  
Author(s):  
Walter J. Muir
Keyword(s):  
Learning Disability ◽  
Gene Dosage ◽  
Genetic Disorders ◽  
Molecular Medicine ◽  
Gene Dosage Effects ◽  
Dosage Effects ◽  
Rapid Changes ◽  
Basic Understanding ◽  
Genetic Mechanisms ◽  
Dynamic Mutations

BackgroundMedicine is rapidly becoming molecular medicine, and little escapes the grasp of modern genetics. Most disorders associated with learning disability have at least a genetic component influencing their expression; in many disorders, disturbances of genetic mechanisms play a pivotal role.AimsDynamic mutations, imprinting mechanisms and gene-dosage effects are explained with reference to genetic disorders that lead to learning disability.MethodA review of recent important studies in the genetics of learning disability.ResultsA host of new genetic connections to conditions associated with learning disability have been made.ConclusionsA basic understanding of these genetic connections is important for all learning disability psychiatrists if they are to follow the rapid changes – already beginning to influence our practice – that hold immense promise for the future.

Selection of a human chromosome 21 enriched YAC sub–library using a chromosome–specific composite probe

1992 ◽  
Vol 1 (4) ◽  
pp. 284-290 ◽  
Author(s):  
Mark T. Ross ◽  
Dean Nižetić ◽  
Catherine Nguyen ◽  
Catherine Knights ◽  
Radost Vatcheva ◽  
...  
Keyword(s):  
Human Chromosome ◽  
Chromosome 21 ◽  
Human Chromosome 21 ◽  
Selection Of
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