Ectopic expression of tethered human follicle-stimulating hormone (hFSH) gene in transgenic mice

Kim, Myoung Ok; Kim, Sung Hyun; Lee, Sang Ryeul; Shin, Mi Jung; Min, Kyeong‐Sik; Lee, Dong Beom; Lee, Sung Won; Kim, Kil Soo; Kim, Sun Jung; Ryoo, Zae Young

doi:10.1007/s11248-006-9031-5

Cited by 11 publications

(10 citation statements)

References 51 publications

(48 reference statements)

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“…In vitro biological activity of hFSH was determined by measuring cyclic-AMP (cAMP) production after exposing hFSH receptor-transfected cells to sterilized milk from each cell line. Milk containing hFSH increased cAMP production in the assay, indicating receptor binding and intracellular signaling, and therefore, biological activity of hFSH (43).…”

Section: Introductionmentioning

confidence: 93%

“…A similar model was later generated that also ectopically expressed FSH in mammary glands, but in this case, the transgene encoded human FSH. In these studies, the investigators used the bovine β-casein gene promoter to specifically express human FSH in mammary glands of transgenic mice (43). Milk was collected from tg-hFSH mice to determine FSH concentration using an enzyme-linked immunosorbent assay (ELISA).…”

Section: Introductionmentioning

confidence: 99%

“…Two mouse lines showed nearly undetectable levels of hFSH, as compared to one line that had high levels (300 mIU/mL). This variation in hFSH concentration in different lines was most likely due to differing transgene copy numbers (43). In vitro biological activity of hFSH was determined by measuring cyclic-AMP (cAMP) production after exposing hFSH receptor-transfected cells to sterilized milk from each cell line.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Gain–of–Function Genetic Models to Study FSH Action

2019

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Follicle–stimulating hormone (FSH) is a pituitary-derived gonadotropin that plays key roles in male and female reproduction. The physiology and biochemistry of FSH have been extensively studied for many years. Beginning in the early 1990s, coincident with advances in the then emerging transgenic animal technology, and continuing till today, several gain-of-function (GOF) models have been developed to understand FSH homeostasis in a physiological context. Our group and others have generated a number of FSH ligand and receptor GOF mouse models. An FSH GOF model when combined with Fshb null mice provides a powerful genetic rescue platform. In this chapter, we discuss different GOF models for FSH synthesis, secretion and action and describe additional novel genetic models that could be developed in the future to further refine the existing models.

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Section: Introductionmentioning

confidence: 93%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Gain–of–Function Genetic Models to Study FSH Action

2019

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“…Human type II collagen was expressed from 1.8-and 3.1-kb bovine β-casein promoters in transgenic mice (Naruse et al, 2006). A 3.7-kb promoter fragment was used to express human follicle-stimulating hormone in transgenic mice (Kim et al, 2007). The reporter gene chloramphenicol acetyl transferase was expressed in transgenic bovine mammary epithelial cells (BMECs) from a 950-bp promoter fragment (Ahn et al, 1995).…”

Section: Introductionmentioning

confidence: 99%

Breeding of transgenic cattle for human coagulation factor IX by a combination of lentiviral system and cloning

Monzani¹,

Sangalli²,

De³

et al. 2013

Genet. Mol. Res.

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ABSTRACT. Recombinant coagulation factor IX must be produced in mammalian cells because FIX synthesis involves translational modifications. Human cell culture-based expression of human coagulation factor IX (hFIX) is expensive, and large-scale production capacity is limited. Transgenic animals may greatly increase the yield of therapeutic proteins and reduce costs. In this study, we used a lentiviral system to obtain transgenic cells and somatic cell nuclear transfer (SCNT) to produce transgenic animals. Lentiviral vectors carrying hFIX driven by 3 bovine β-casein promoters were constructed. Bovine epithelial mammary cells were transduced by lentivirus, selected with blasticidin, plated on extracellular matrix, and induced by lactogenic hormones; promoter activity was evaluated by quantitative PCR. Transcriptional activity of the 5.335-kb promoter was 6-fold higher than the 3.392-and 4.279-kb promoters, which did not significantly differ. Transgenic bovine fibroblasts were transduced with lentivirus carrying the 5.335-kb promoter and used as donor cells for SCNT. Cloned transgenic embryo production yielded development rates of 28.4%, similar to previous reports on cloned non-transgenic embryos. The embryos were transferred to recipient cows (N = 21) and 2 births of cloned transgenic cattle were obtained. These results suggest combination of the lentiviral system and cloning may be a good strategy for production of transgenic cattle.

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“…However, there are some disadvantages in traditional transgenic research, including limited regulatory elements, random insertion and repeated nature of the multiple copy number integration, which results in low level expression, even with ectopic expression of recombinant proteins (al-Shawi et al, 1990;Clark et al, 1994;Dobie et al, 1996;Kim et al, 2007). In addition, the random integration of exogenous genes into the host genome may inadvertently affect the expression of endogenous genes by disabling them or activating them at an inappropriate time, which is dangerous for the transgenic animal (Seggewiss et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

Targeting the human lysozyme gene on bovine αs1-casein gene locus in fibroblasts

Cheng¹

2011

Afr. J. Biotechnol.

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Targeting an exogenous gene into a favorable gene locus and for expression under endogenous regulators is an ideal method in mammary gland bioreactor research. For this purpose, a gene targeting vector was constructed to targeting the human lysozyme gene on bovine αs1-casein gene locus. In this case, the expression of human lysozyme could be regulated by the endogenous cis-element of αs1casein gene in bovine mammary glands. In order to analyze the bioactivity of the vector, the targeting vector was stably transfected and randomly integrated into mouse mammary epithelial cells. Reversetranscription-polymerase chain reaction (RT-PCR) and western blot results showed that, the bovine αs1-casein promoter in the 5' arm was able to direct the efficient expression and secretion of human lysozyme in mammary epithelial cells. Turbidimetric assay showed that the antibacterial activity of lysozyme in transfected cells culture medium was 180 U/ml. To obtain the gene targeted cells line, bovine fetal fibroblasts were isolated and transfected with linear targeting vector (21.9 kb) using nucleofector device, which the transfection rate was about 25%. After seven rounds of independent cell transfection, a total of 8 × 10 7 cells were transfected, 118 colonies were expanded and analyzed by PCR, but none were found to be targeted. However, the targeted events were detected in the mixed cells which did not formed obvious colonies in five 10 cm dishes. Thus, these results indicate that, the 8.2 kb exogenous genes could be site specifically integrated into the transcriptionally silent αs1-casein gene locus in fibroblasts, but the unfavorable chromatin structure in such loci may have a disadvantage to targeted colonies formation in expansion stage. We suggest that minimizing the length of in vitro culture time and relax selection as soon as colonies become evident might prevent such loss of targeted cells.

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Ectopic expression of tethered human follicle-stimulating hormone (hFSH) gene in transgenic mice

Cited by 11 publications

References 51 publications

Gain–of–Function Genetic Models to Study FSH Action

Gain–of–Function Genetic Models to Study FSH Action

Breeding of transgenic cattle for human coagulation factor IX by a combination of lentiviral system and cloning

Targeting the human lysozyme gene on bovine αs1-casein gene locus in fibroblasts

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