Expression of eukaryotic recombinant proteins and deriving them from the milk of transgenic animals

Khodarovich, Yu. M.; Gol'dman, I. L.; Садчикова, Е.Р.; Georgiev, Pavel

doi:10.1134/s0003683813090020

Cited by 6 publications

(15 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…To the best of our knowledge, the present study is the first to report the expression level and stability of rhPA in the milk produced by homozygote transgenic rabbits (4). A novel PcL25/rhPA transgenic rabbit strain (K29) was established, and high expression levels of rhPA was achieved in the homozygous transgenic rabbit (5,6).…”

Section: Discussionmentioning

confidence: 89%

“…Recombinant proteins are the main biopharmaceutical agents (1)(2)(3)(4), and the reduction of primary cost of recombinant proteins and improvement of their quality by selecting novel efficient expression systems has received increasing attention (1)(2)(3)(4). The production of recombinant proteins through the use of bacterial hosts, mammalian cells, yeast, viruses, transgenic plants or other expression systems has numerous disadvantages, including poor biological activity, high cost and poor protein post-translation modification among other (1)(2)(3)(4). Recombinant proteins expressed in the mammary gland of transgenic animals have become a research focus as bioreactors due to their high expression, low cost, easy purification process and similar protein modification to that in the human body (4)(5)(6)(7)(8)(9)(10)(11).…”

Section: Introductionmentioning

confidence: 99%

“…The production of recombinant proteins through the use of bacterial hosts, mammalian cells, yeast, viruses, transgenic plants or other expression systems has numerous disadvantages, including poor biological activity, high cost and poor protein post-translation modification among other (1)(2)(3)(4). Recombinant proteins expressed in the mammary gland of transgenic animals have become a research focus as bioreactors due to their high expression, low cost, easy purification process and similar protein modification to that in the human body (4)(5)(6)(7)(8)(9)(10)(11). Therefore, the animal mammary gland is currently the most ideal expression system for the production of recombinant proteins.…”

Section: Introductionmentioning

confidence: 99%

“…Various recombinant proteins, such as ATryn, hAAT, recombinant human protein c, superoxide dismutase (SOd) and hormones, have been generated in the milk of transgenic animals (8)(9)(10)(11)(12). However, although the first production of recombinant proteins through the milk of transgenic animal for therapeutic use was reported 30 years ago, the recombinant proteins expression regulatory mechanism in the milk of these animals remains unclear and only a few recombinant proteins have been successfully produced through the transgenic animal mammary gland (4,13).…”

Section: Introductionmentioning

confidence: 99%

“…The expression of recombinant proteins in the mammary gland of transgenic animals is a complex process affected by various factors, including the genetic structure, gene copy number and their regulatory elements, gene integration site, promoter and methylation (3,4). At present, the expression level and stability of recombinant proteins in transgenic animals are not sufficient to allow for larger-scale production of the desired proteins (3,4).…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

Efficient increase of the novel recombinant human plasminogen activator expression level and stability through the use of homozygote transgenic rabbits

Jiang

Zhang

et al. 2018

Int J Mol Med

View full text Add to dashboard Cite

Expression efficacy of recombinant protein in current expression systems is generally low. Therefore, the expression levels of recombinant proteins in the breast milk of transgenic animals are typically low. In view of this, the present study aimed to construct homozygous transgenic rabbits with a high expression level of recombinant human plasminogen activator (rhPA) during the entire lactation period. Homozygous transgenic rabbits were obtained using an effective rhPA mammary‑specific expression vector PCL25/rhPA. The expression level and thrombolytic ability of rhPA in the milk of both homozygous and hemizygous rabbits were detected by enzyme‑linked immunosorbent and fibrin agarose plate assays. It was observed that the expression of rhPA was constant during the entire lactation period in homozygous rabbits, while the expression of rhPA declined slowly in hemizygote rhPA transgenic rabbits during the lactation period. In addition, the expression of rhPA in homozygous transgenic rabbit was ~950 µg/ml, which was markedly higher in comparison with that in hemizygote rabbits. Furthermore, increased gene copy number was observed to increase the expression level of rhPA at the same integration vector.

show abstract

Section: Discussionmentioning

confidence: 89%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Efficient increase of the novel recombinant human plasminogen activator expression level and stability through the use of homozygote transgenic rabbits

Jiang

Zhang

et al. 2018

Int J Mol Med

View full text Add to dashboard Cite

show abstract

Identification, purification, and pharmacological activity analysis of Desmodus rotundus salivary plasminogen activator alpha1 (DSPAα1) expressed in transgenic rabbit mammary glands

et al. 2022

View full text Add to dashboard Cite

A novel recombinant human plasminogen activator: Efficient expression and hereditary stability in transgenic goats and in vitro thrombolytic bioactivity in the milk of transgenic goats

Lü

Zhang

et al. 2018

PLoS ONE

View full text Add to dashboard Cite

BackgroundThromboses is a rapidly growing medical problem worldwide. Low-cost, high-scale production of thrombotic drugs is needed to meet the demand. The production of biomolecules in transgenic animals might help address this issue. To our knowledge, the expression of recombinant human plasminogen activator (rhPA) in goat mammary glands has never been reported before.MethodsWe constructed a mammary gland–specific expression vector, BLC14/rhPA, which encodes only the essential K2 fibrin-binding and P domains of wild-type tPA (deletion mutant of tPA lacking the F, E, and K1 domains), along with the goat β-lactoglobulin gene signal peptide-coding sequence. The mammary gland–specific expression vector BLC14/rhPA was transfected into goat fetal fibroblast cells by electroporation. After selection for 3 weeks by G418, stably transfected cell colonies were obtained. PCR analysis results indicated that 24 of the resistant clones were transgenic cell lines; of these, 8 lines were selected as the donor cells. The positive cells were starved for 72 h with DMEM/F12 medium containing 0.5% FBS and were then used as do. Finally, 256 reconstructed oocytes were transferred into 26 recipients, and 7 of them became pregnant (pregnancy rate, 26.9%). Two kids were obtained (BP21 and BP22). PCR analysis confirmed that both were transgenic goats. To analyze the heredity of the rhPA expressed in BP21 F0 and F1 transgenic goats, the F0 transgenic goat BP21 was mated with a normal male goat to generate an F1 transgenic goat. Enucleated metaphase II (MII) oocytes and positive donor cells were used to reconstruct embryos, which were transplanted into the oviducts of the recipients.ResultsWestern blot results showed a specific 39 kDa band. The rhPA expression level in transgenic goat whey was about 78.32 μg/mL by ELISA. Results of ELISA and the in vitro thrombolysis test (FAPA) showed that specific activity of the rhPA in the milk of F0 and F1 transgenic goats was 13.3 times higher than that of the reteplase reference material.ConclusionThus, we demonstrated that BLC14/rhPA was reasonably effective for expression in the mammary glands of transgenic goats, and was stably inherited by the offspring. This study provides the basis for the large-scale production of biological pharmaceuticals in transgenic animals. The expression of biopharmaceuticals by transgenic animals can be used for pharmacological research and bioactive analysis, and transgenic goats were demonstrated to be promising animals for the large-scale production of thrombolytic biopharmaceuticals.

show abstract

Expression of eukaryotic recombinant proteins and deriving them from the milk of transgenic animals

Cited by 6 publications

References 33 publications

Efficient increase of the novel recombinant human plasminogen activator expression level and stability through the use of homozygote transgenic rabbits

Efficient increase of the novel recombinant human plasminogen activator expression level and stability through the use of homozygote transgenic rabbits

Identification, purification, and pharmacological activity analysis of Desmodus rotundus salivary plasminogen activator alpha1 (DSPAα1) expressed in transgenic rabbit mammary glands

A novel recombinant human plasminogen activator: Efficient expression and hereditary stability in transgenic goats and in vitro thrombolytic bioactivity in the milk of transgenic goats

Contact Info

Product

Resources

About