Expression of a foreign gene in a line of transgenic mice is modulated by a chromosomal position effect.

Al‐Shawi, Raya; Kinnaird, Jane; Burke, J; Bishop, John O.

doi:10.1128/mcb.10.3.1192

Cited by 158 publications

(68 citation statements)

References 44 publications

(40 reference statements)

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“…Although this approach is potentially applicable to any mammalian cell line, the HEK293 cell line was chosen because of its ease of transfection, handling, and rapid cell growth. A robust system for coexpressing IgG and AID in mammalian cells was created using episomal vectors expressing the heavy chain (HC), light chain (LC), and AID, each also encoding a separate antibiotic selectable marker (20,21). To display immunoglobulin on the surface of these cells, full-length human IgG1 HC was modified by addition of a C-terminal transmembrane domain (22).…”

Section: Resultsmentioning

confidence: 99%

Coupling mammalian cell surface display with somatic hypermutation for the discovery and maturation of human antibodies

Bowers

Horlick

Neben

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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A novel approach has been developed for the isolation and maturation of human antibodies that replicates key features of the adaptive immune system by coupling in vitro somatic hypermutation (SHM) with mammalian cell display. SHM is dependent on the action of the B cell specific enzyme, activation-induced cytidine deaminase (AID), and can be replicated in non-B cells through expression of recombinant AID. A library of human antibodies, based on germline V-gene segments with recombined human D J regions was used to isolate low-affinity antibodies to human β nerve growth factor (hβNGF). These antibodies, initially naïve to SHM, were subjected to AID-directed SHM in vitro and selected using the same mammalian cell display system, as illustrated by the maturation of one of the antibodies to low pM K D . This approach overcomes many of the previous limitations of mammalian cell display, enabling direct selection and maturation of antibodies as full-length, glycosylated IgGs. affinity maturation | mammalian display

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Section: Resultsmentioning

confidence: 99%

Coupling mammalian cell surface display with somatic hypermutation for the discovery and maturation of human antibodies

Bowers

Horlick

Neben

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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“…4) (Table II). C, Southern analyses are as described for B. Fragments in lanes 1,6,7, and 12 are larger than expected (arrow), indicating the loss of either or both restriction enzyme sites. Duplicate mice of lines A4/2-17 and A2/4 -29 were analyzed in lanes 8 -11.…”

Section: Potential Mechanisms Of Blocking By the Alu2 Fragment-mentioning

confidence: 99%

“…Position effects may result from the influence of regulatory elements of the flanking DNA, from repression by nearby heterochromatin (1-3), or possibly from transcriptional interference of one gene on another (4). In vertebrates, position effects are common in transgenic mice and after chromosomal translocations (3,5,6).…”

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confidence: 99%

An Alu Element from the K18 Gene Confers Position-independent Expression in Transgenic Mice

Willoughby

Vilalta

Oshima

2000

Journal of Biological Chemistry

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We have identified a 323-base pair fragment of the 5-flanking sequence of the K18 gene, which confers position-independent and copy number-dependent expression on two heterologous transgenes. This fragment is composed primarily of an Alu repetitive element. Its activity in mice is correlated with its RNA polymerase III promoter activity and its orientation-dependent ability to inhibit potential transcriptional interference in a transfection assay. However, the activity of the Alu element is not correlated with its enhancer blocking activity, a characteristic of insulator elements. In addition, this Alu element did not block the suppressive effect of co-injecting mouse ␣ satellite DNA with the transgene. This Alu element is likely responsible for at least part of the protective effects of the sequences flanking the K18. These results suggest that transcriptionally active Alu elements may eliminate transcriptional interference of neighboring genes. This Alu element is one component of the locus control region associated with the K18 gene. Other Alu repetitive elements may also function to define regulatory domains.

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“…A small subpopulation of the interneurons, granular and periglomerular cells, is also stained in the OB, which might be of the correct type, as these cells are considered to be GABAergic (Mugnaini et al 1984). The incorrect expression of the transgene, frequently found in transgenic mice, most probably results from site of integration effects (Al-Shawi et al 1990;Weis et al 1991;Bessis et al, 1995). The purpose of this study was to find a reliable method for direct identification of the synaptic interactions of well-defined neuronal cell types, ectopically labeled, ␤ -galactosidasepositive output neurons in particular.…”

Section: Discussionmentioning

confidence: 99%

Visualization of β-Galactosidase by Enzyme and Immunohistochemistry in the Olfactory Bulb of Transgenic Mice Carrying the LacZ Transgene

Sekerková

Katarova

Joó

et al. 1997

J Histochem Cytochem.

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SUMMARYIn the olfactory bulb (OB) of a transgenic mouse line that carries the bacterial LacZ gene under the control of the 5 -regulatory region of the GAD67 gene, expression of the ␤ -galactosidase was confined almost exclusively to the non-GABAergic mitral and tufted cells. By light microscopy, enzyme histochemistry showed strong staining in the cell bodies and faint diffuse staining in the axons and dendrites. With immunohistochemistry for ␤ -galactosidase the entire cytoplasm, including the axons and dendrites, was strongly stained. By electron microscopy, ␤ -galactosidase enzyme histochemistry resulted in a submicroscopic reaction product that was diffusely distributed in the cytoplasm of neurons. In addition, large deposits of the reaction product were also seen attached to the cytoplasmic side of the membranes. In contrast, when the intracellular localization of ␤ -galactosidase was determined by immunohistochemistry, homogeneous cytoplasmic staining was obtained that filled the entire cytoplasm including the terminal dendrites and fine axons. Therefore, synaptic contacts of the ␤ -galactosidase-positive output neurons with other ␤ -galactosidase-negative neuronal cells were readily recognized in the OB. As we demonstrated, transgenic mouse lines expressing the LacZ reporter gene in a well-defined neuronal subpopulation can be used to follow ␤ -galactosidase-positive neurons and to directly identify their synaptic connections. (J Histochem Cytochem 45:1147-1155, 1997)

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Expression of a foreign gene in a line of transgenic mice is modulated by a chromosomal position effect.

Cited by 158 publications

References 44 publications

Coupling mammalian cell surface display with somatic hypermutation for the discovery and maturation of human antibodies

Coupling mammalian cell surface display with somatic hypermutation for the discovery and maturation of human antibodies

An Alu Element from the K18 Gene Confers Position-independent Expression in Transgenic Mice

Visualization of β-Galactosidase by Enzyme and Immunohistochemistry in the Olfactory Bulb of Transgenic Mice Carrying the LacZ Transgene

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