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1

Noninvasive fluorescent screening of microinjected bovine embryos to predict transgene integration

100%
Rosochacki S. J., Kozikova L., Korwin-Kossakowski M., Matejczyk M., Poloszynowicz J., Duszewska A. M.
Folia Biologica
|
2003
|
vol. 51
|
issue 1-2
97-104
EN
Most transgenic domestic animals are generated by direct microinjection of DNA fragments into zygote pronuclei. It has generally been assumed that the majority of integration events should occur prior to the first round of chromosomal DNA replication. The aim of this study was to investigate the expression of GFP in bovine preimplantation embryos by using a gfp reporter gene consisting of chicken beta-actin promoter, the CMV-IE enhancer, gfp cDNA (EGFP) (732 bp) and rabbit beta-globin polyadenylation sequences. In five experiments 302 bovine zygotes were injected while 75 served as a control. The fluorescence intensity was detected at 72 and 168 h following fertilization in bovine embryos injected with 3 ng/mu l in experiments 1-3, and injected with 5 ng/mu l in experiments 4-5. Eight embryos were considered as expressing green fluorescence protein; 2 of them were 100% fluorescent after microinjection of a higher dose of the DNA; one was 75%, two - 50%, and three 25% transgenic. The mosaicism was assumed to be at 75%. The results indicated that the fluorescence could be analyzed at any time of bovine embryo development. It was therefore concluded, that chicken -actin promoter together with the CMV-IE enhancer would confer a strong expression of the gfp reporter gene in preimplantation bovine embryos. Therefore, using GFP that could be simply detected in live bovine (transgenic) embryos would be very promising in establishing transgenic lines of domestic animals producing in their fluids human therapeutic proteins.
2

Characterization and application of embryonic stem cells ESC in animal biotechnology

100%
Lechniak D.
Biotechnologia
|
1993
|
issue 2
55-62
EN
The micromanipulations of embryos and gametes are important tasks in animal biotechnology. These techniques can be used in animal cloning and transgenesis. For achieving these goals, embryonic stem cells are of principial interest. In vitro cultures of ESC can be derived from whole embryos or isolated inner cell masses (ICM). The attraction of ESC to animal biotechnological research is their pluripotency. ESC are able to contribute to all tissues, including germ lines. To obtain transgenic animals, ESC can be genetically transformed prior to the introduction to embryos. Present status of knowledge of ESC and their possible usefulness for genetic improvement of farm animals are discussed.
3

The expression of WAP-Fuc gene in genotypes of transgenic farm animals ? effectiveness of transgenesis with the use of DNA microinjection

100%
Jura J., Smorag Z., Gajda B., Kareta W., Kopchick J. J., Kelder B., Prieto P. A.
|
2003
|
issue 1
216-222
EN
There are two principal applications of transgenic animals. Best known and most advanced application is to use transgenic animals (called bioreactors or molecular farms) for the production of various proteins or biopolymers of medical significance. The second application concerns efforts to improve the productivity traits of breeding animals. We have worked out methods which allow somatic cloning of mammals and gene knockout methods. These methods have been developing very rapidly in recent years and their efficiency will soon be improved to the extent that they will become profitable. For the time being, DNA microinjection with all its disadvantages, remains the principal method of producing transgenic bioreactors. In this paper, the effectiveness of production of transgenic rabbits, goats and pigs with the use of WAP-Fuc gene construct is presented.
4

Production of therapeutic proteins in the semen of transgenic animals

100%
Lipinski D., Juzwa W., Zeyland J., Slomski R.
Biotechnologia
|
2006
|
issue 1
44-52
EN
The expression of the recombinant proteins by transgenic animals represents an opportunity to achieve cost-effective, large-scale production of a wide variety of therapeutics. Among transgenic animal production systems, the transgenic mammary gland is the most advanced. However, the production of proteins in milk is limited by a relatively long interval from birth to first lactation encountered with domestic livestock, the discontinuous nature of the lactation cycle and the substantial time and material investments required to produce transgenic dairy animals. The semen of transgenic boar represents an alternative platform for the production of therapeutic proteins. The expression of such proteins in the male accessory glands, particularly in the seminal vesicle epithelium can be controlled by gene regulatory sequences specific to these tissues. In this review, we consider the possibility of using such regulatory sequences to drive the production of foreign proteins into the seminal fluids of transgenic animals. Application of this technology to pigs which can be ejaculated 2-3 times per week (200-300 ml per ejaculate), could lead to the annual production of several grams of recombinant protein.
5

Biotechnology in potato breeding

100%
Marczewski W.
Biotechnologia
|
2008
|
issue 2
20-26
EN
Potato has been known in Europe for the past 400 years. Potato breeding began after the global Phytophthora infestans epidemics in 1840 and 1841. The first breeding attempts were to increase the P. infestans resistance in cultivated potato. In the 21st century, new potato cultivars have been evaluated for more than 50 traits, both for disease and pest resistance, and significant tuber traits. Modern potato breeding requires biotechnology. Cell and tissue culture, genetic engineering and DNA-based molecular markers are the most promising areas of molecular biology for potato breeding. However, there are few reports indicating the actual use of biotechnology in potato breeding programs. Cisgenesis and DNA markers that reside within resistance genes or physically close to them and identified in a fully automated system are a chance for the progress in the future.
6

Human antibody expression in transgenic mice

100%
Bruggemann M.
Archivum Immunologiae et Therapiae Experimentalis
|
2001
|
vol. 49
|
issue 3
203-208
EN
Human antibody repertoires can be created in transgenic mice following the introduction of human immunoglobulin heavy and light chain genes in their germline configuration. Transgene constructs or transloci have been obtained by plasmid assembly, cloning in yeast artificial chromosomes, and the use of chromosome fragments. Translocus integration and maintenance in transgenic mouse strains has been achieved by pronuclear DNA injection into oocytes and various transfection methods using embryonic stem cells. The human DNA segments rearrange faithfully in the mouse and produce extensive V(D)J combinations. Specific human monoclonal antibodies of high affinity for use in therapeutic applications have been produced from these translocus mice.
7

Expression of microinjected reporter gene lacZ during first cleavages of rabbit embryos

100%
Rosochacki S. J., Strzelecka M., Kozikova L., Matejczyk M., Oblap R., Poloszynowicz J., Zwierzchowski L.
Folia Biologica
|
2002
|
vol. 50
|
issue 1-2
61-67
EN
The growing use of reporter genes in a model transgenic system has been a fundamental approach of biology, but the strategy of transgenic embryo selection prior to transfer to foster mothers may greately increase the efficiency of transgenic livestock production. This study was conducted to assess the possibility of beta -galactosidase (beta -gal)-labeled transgenic rabbit embryo production. Rabbit zygotes were obtained from superovulated females after mating. Zygotes were microinjected into male pronuclei with pCMV-lacZ or SV40-lacZ constructs; while some embryos were coinjected with the scaffold attachment sequences - SAR. Embryos from control non-injected and microinjected groups were cultured in vitro. After 24, 48, 72, or 96 h of culture the embryos were stained with X-gal for beta -galactosidase. Transgenic embryos produced by pronuclear injection showed a discrete pattern of beta -galactosidase expression. The percentage of transgenesis with pCMV-lacZ alone was 1.5, but with SAR sequences it increased to 4.2. In the case of SV40-lacZ construct, the efficiency of transgenesis was 2.3% and 4.1%, respectively. The mosaicism was 66.7% for all embryos injected with both constructs with or without SAR. The highest numbers of 100%-transgenic (non-mosaic) embryos were found in the group co-injected with SV40-lacZ and SAR. Transgenesis was seen as early as 24 h after injection, in four-cell embryos. Most of the microinjected embryos showed delayed development as compared with control. It was concluded that lacZ may serve as a reliable reporter for early transgenic embryo selection in order to produce transgenic animals.
8

Modifications of animal tissues by way of genetic engeneering and their possible significance in xenotransplantation

100%
Grzybowski G.
Biotechnologia
|
1998
|
issue 1
95-104
EN
The paper presents the problems connected with the modification of animal tissues through transgenesis. Special attention has been given to the xenogenic heart transplant between pigs and human beings. The heart of pigs is a highly discordant graft both for primates and human, which is hyperacutely rejected already several minutes after operation. Graft rejection is mainly due to the presence of natural xenoreactive antibodies which recognize the pigs' tissue epitop Gala1-3Gal (known as the Gal antigene), as well as to the quick activation of the complement in the recipient. In 1995 transgenic pigs with human genes responsible for complement CD55 (DAF - decay accelerating factor) and CD59 factor (membrane inhibitor of reactive lysis) were obtained. Organs of such animals, grafted into baboons, were resistant to complement-related rejection. The possibilities of modifying animal tissues by way of genetic manipulation are still limited and total 'humanzation' of pigs heart as regards the histocompatibility antigens is not yet feasible. For this reason the fulfilment of the idea of xenografting between pigs and human beings is still remote.
9

Transgenic farm animals - efectiveness of transgenesis via microinjection

100%
Jura J.
Biotechnologia
|
1995
|
issue 3
20-32
EN
The gene construct and the kind of its promotor have the major influence on the effectiveness of transgenesis via microinjection.A new succesful transgenesis technique is described.
10

Animal transgenesis: state of the art and applications

100%
Melo E. O., Canavessi A. M. O., Franco M. M., Rumpf R.
Journal of Applied Genetics
|
2007
|
vol. 48
|
issue 3
47-61
EN
There is a constant expectation for fast improvement of livestock production and human health care products. The advent of DNA recombinant technology and the possibility of gene transfer between organisms of distinct species, or even distinct phylogenic kingdoms, has opened a wide range of possibilities. Nowadays we can produce human insulin in bacteria or human coagulation factors in cattle milk. The recent advances in gene transfer, animal cloning, and assisted reproductive techniques have partly fulfilled the expectation in the field of livestock transgenesis. This paper reviews the recent advances and applications of transgenesis in livestock and their derivative products. At first, the state of art and the techniques that enhance the efficiency of livestock transgenesis are presented. The consequent reduction in the cost and time necessary to reach a final product has enabled the multiplication of transgenic prototypes around the world. We also analyze here some emerging applications of livestock transgenesis in the field of pharmacology, meat and dairy industry, xenotransplantation, and human disease modeling. Finally, some bioethical and commercial concerns raised by the transgenesis applications are discussed.
11

Loop formation by the transgene WAP:6xHishGH in transgenic rabbit fibroblasts, revealed by fluorescence in situ hybridization to nuclear halos

100%
Michalak E., Lipinski D., Slomski R.
|
|
issue 3
247-249
EN
Using fluorescence in situ hybridization (FISH) to somatic nuclear halos from transgenic rabbits WAP:6xHishGH, we present evidence for stability of transgenesis at the chromatin level. FISH performed on fibroblasts from a homozygous individual showed 2 independent loops from both chromosomes of pair 7. On a heterozygous individual, FISH detected a single loop. According to the concept of chromatin loops and their influence on gene expression, this shows that the human growth hormone transgene, which was actively expressed in mammary gland under the influence of the tissue-specific promoter, was inactive in examined skin fibroblasts.
12

Animal transgenesis: state of the art and applications

100%
Melo E. O., Canavessi A. M. O., Franco M. M., Rumpf R.
Journal of Applied Genetics
|
2007
|
vol. 48
|
issue 1
47-61
EN
There is a constant expectation for fast improvement of livestock production and human health care products. The advent of DNA recombinant technology and the possibility of gene transfer between organisms of distinct species, or even distinct phylogenic kingdoms, has opened a wide range of possibilities. Nowadays we can produce human insulin in bacteria or human coagulation factors in cattle milk. The recent advances in gene transfer, animal cloning, and assisted reproductive techniques have partly fulfilled the expectation in the field of livestock transgenesis. This paper reviews the recent advances and applications of transgenesis in livestock and their derivative products. At first, the state of art and the techniques that enhance the efficiency of livestock transgenesis are presented. The consequent reduction in the cost and time necessary to reach a final product has enabled the multiplication of transgenic prototypes around the world. We also analyze here some emerging applications of livestock transgenesis in the field of pharmacology, meat and dairy industry, xenotransplantation, and human disease modeling. Finally, some bioethical and commercial concerns raised by the transgenesis applications are discussed.
13

Production of the human hemoglobin from animals modified by genetic engineering

88%
Grzybowski G.
Biotechnologia
|
1998
|
issue 1
83-94
EN
Medical and economic arguments for the investigation of methods of producing blood substitutes are presented. The paper presents the current state of investigations aimed at the production of human hemoglobin from animals modified by genetic engineering methods. Certain problems related to medicine and public perception are presented in connection with the use of 'transgenic' hemoglobin as blood substitute. Swine is at present the only species among farm animals which is used in research on the production of human hemoglobin. Animals were obtained in which 'transgenic' hemoglobin accounted for 54% of the total hemoglobin in the organism. When human hemoglobin accounted for 24% of the total, and 30% was hybrid (human/swine), the animals were in perfect health, fit for reproduction and their progeny demonstrated the same transgene expression. It is assumed that human hemoglobin, extracted from the blood of transgenic swine, will be the first commercial product used in medicine and obtained through transgenesis of domestic animals.
14

The role of domestic pig in somatic cell cloning and transgenesis

88%
Skrzyszowska M., Samiec M.
Biotechnologia
|
2006
|
issue 1
53-67
EN
A stimulus for development of the studies on pig somatic cell cloning, especially in recent years, was above all the possibility of its practical application for production of transgenic piglets using in vitro transfected nuclear donor cells and multiplication of genetically-engineered sows and boars generated so far, on the grounds of important implications for biomedicine, pharmacy and agriculture. However, effective pig somatic cell nuclear transfer, avoiding the sexual reproduction pathway, creates a possibility of providing numerous monogenetic and monosexual offspring derived not only from genetically-transformed individuals, but also from adult (postpubertal) animals of high genetic merit. Generation of cloned transgenic pigs for biomedical purposes to obtain recombinant xenogeneic proteins or organs suitable in xenotransplantology, or to create cell (gene) therapy foundations for a number of serious monogenic diseases that induce heritable (congenital) developmental anomalies, is perceived as a service to humanity.
15

Development of the studies on somatic cell cloning in goats

88%
Skrzyszowska M., Samiec M.
Biotechnologia
|
2006
|
issue 1
133-150
EN
Domestic goat as a species with a relatively great biodiversity of dairy breeds, which possess high genetic merit and yield of milk production, can be a valuable tool for embryo gene engineering. This involves the generation of transgenic specients, providing with xenogeneic (human) recombinant proteins (i.e. biopharmaceuticals), not only by the standard zygote intrapronuclear microinjection of gene constructs, but above all with the use of somatic cell cloning technology.
16

Present-day and future state of art of reconstruction of the facial skeleton

88%
Zapala J., Wyszynska-Pawelec G.
Biotechnologia
|
2006
|
issue 1
110-124
EN
In this paper, present-day techniques used in reconstructive surgery of the facial skeleton are presented, including indications for autogenous, free and vascularized bone grafts as well as homogenous grafts, implants and biomaterials. The main directions of the development of reconstructive surgery are discussed. The preliminary results of research on xenogenous bone grafts harvested from transgenic pigs are presented as well as experimental model for New Zealand white rabbit and transgenic pig.
17

Reproduction biotechnology of farm animals

75%
Smorag Z.
Biotechnologia
|
2002
|
issue 4
30-40
EN
This paper presents the most important research and application of biotechnology of farm animals reproduction: semen sexing, embryo in vitro production, embryonic and somatic cloning and transgenesis.
18

Production of pigs used in xenotransplantation

75%
Jura J., Slomski R., Smorag Z., Gajda B., Wieczorek J., Lipinski D., Kalak R., Juzwa W., Zbyland J.
Biotechnologia
|
2006
|
issue 1
151-158
EN
There are four main trends in the use of transgenic animals. One of the most interesting is the use of transgenic animals as the donors of tissue or organs suitable for transplantation in human ? xenotransplantation. This field of research has been undergoing intensive and increasing study during the past few years, and some encouraging progress is being made. A pig has been identified as the most suitable donor animal. The aim of the presented experiment was to produce transgenic pigs which tissues and organs could be used for the xenotransplantation needs. To target the goal, a competitive gene construct coding the same substrate as the endogenous enzyme of organ donor was introduced. In effect, transgenic boar was produced with confirmed integration of human a1,2 fucosyltransferase gene. Also, F1 generation of transgenic pigs was generated to preserve ongoing needs of preliminary research of xenotransplantation project.
19

Achievements in biotechnology of animal reproduction over the past 20 years - case studies

63%
Smorag Z.
|
EN
Animal reproduction biotechnology is an area of fast development with possibilities of practical applications not only in breeding, but in pharmacy and biomedicine as well. Its growth has been achieved thanks to the results of last decades of the previous century in embryology, endocrinology, and molecular biology. The paper contains very brief description of the achievements of NRIAP Department of Animal Reproduction Biotechnology during the last 20 years in mammalian sex regulation, cryobiology of gametes and embryos, in vitro production of embryos, cloning and animal transgenesis.
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