scholarly journals Strategy for modification of the bovine beta-lactoglobulin geneusing components of the CRISPR/Cas9 system in plasmid form

2020 ◽  
Vol 10 (5) ◽  
pp. 211-216
Author(s):  
E.M. Koloskova ◽  
V.A. Ezerskiy ◽  
K.S. Ostrenko
Keyword(s):  
Protein Gene ◽  
Fluorescent Protein ◽  
Specific Activity ◽  
Green Fluorescent Protein Gene ◽  
Regulatory Sequences ◽  
Reporter Protein ◽  
Genetic Construct ◽  
System Components ◽  
Beta Lactoglobulin

Using on-line programs, sites were selected for obtaining double-stranded breaks in the BLG gene of cattle. The strategy for making double-stranded cuts in the BLG gene was developed taking into account the polymorphic variant of the gene (A-allele): DNA was isolated from bovine sperm used for fertilization of cow eggs in vitro. Four pX330 plasmids encoding Cas9 endonuclease and gRNAs specific to the selected BLG gene sequences were obtained. A strategy was developed for analyzing possible genetic modifications resulting from the operation of the CRISPR/Cas9 system components and the genetic construct microinjected into zygotes (NHEJ, HDR). The pBLGcmvEGFP plasmid containing the green fluorescent protein gene under the cytomegalovirus promoter was proposed as a model genetic construct for replacing the BLG gene. The use of a plasmid containing the reporter protein gene under its own regulatory sequences, flanked by homology arms to the beta-lactoglobulin gene, can be useful for evaluating the effectiveness of site-specific activity of the CRISPR/Cas9 system components in vitro.

Efficient Knockout of Transplanted Green Fluorescent Protein Gene in Medaka Using TALENs

2014 ◽  
Vol 16 (6) ◽  
pp. 674-683 ◽  
Author(s):  
Chao Qiu ◽  
Bin Cheng ◽  
Yunsheng Zhang ◽  
Rong Huang ◽  
Lanjie Liao ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Protein Gene ◽  
Fluorescent Protein ◽  
Green Fluorescent Protein Gene ◽  
Green Fluorescent

O4-01-05: Profile and regulation of APOE expression in central nervous system in mice with targeting of green fluorescent protein gene to the APOE locus

2006 ◽  
Vol 2 ◽  
pp. S75-S76
Author(s):  
Qin Xu ◽  
Aubrey Bernardo ◽  
David Walker ◽  
Tiffany Kanegawa ◽  
Robert W. Mahley ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Green Fluorescent Protein ◽  
Protein Gene ◽  
Fluorescent Protein ◽  
Green Fluorescent Protein Gene ◽  
Apoe Locus ◽  
Green Fluorescent ◽  
Apoe Expression

scholarly journals Quantitative Analysis of Tetracycline-Inducible Expression of the Green Fluorescent Protein Gene in Transgenic Chickens

2012 ◽  
Vol 58 (6) ◽  
pp. 672-677 ◽  
Author(s):  
Bon Chul KOO ◽  
Mo Sun KWON ◽  
Ji Yeol ROH ◽  
Minjee KIM ◽  
Jin-Hoi KIM ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Quantitative Analysis ◽  
Protein Gene ◽  
Fluorescent Protein ◽  
Inducible Expression ◽  
Green Fluorescent Protein Gene ◽  
Green Fluorescent ◽  
Transgenic Chickens

183 DEVELOPMENT OF PORCINE EMBRYOS MICROINJECTED WITH PORCINE EMBRYONIC GERM CELLS

2006 ◽  
Vol 18 (2) ◽  
pp. 199
Author(s):  
C.-H. Park ◽  
S.-G. Lee ◽  
D.-H. Choi ◽  
M.-G. Kim ◽  
C. K. Lee
Keyword(s):  
Germ Cells ◽  
Fluorescent Protein ◽  
Inner Cell Mass ◽  
Es Cells ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Green Fluorescent Protein Gene ◽  
Cleavage Stage ◽  
Allocation Pattern

Embryonic germ (EG) cells, derived from primordial germ cells in the developing fetus, are similar to embryonic stem (ES) cells in terms of expression pattern of undifferentiated markers and their ability to colonize both the somatic and the germ cell lines following injection into a host blastocyst, which has been proven in mouse. Several studies using porcine EG cells have shown that it is possible to produce somatic chimeras after blastocyst injection. However, not only was the degree of reported chimerism low, but also there has been no report about the fate of injected EG cells in porcine blastocysts. This study was designed to observe the distribution pattern of porcine EG cells in chimeric blastocyst after injection into cleavage-stage porcine embryos. To ascertain development of microinjected porcine embryos with EG cells, 10 to 15 EG cells were injected into cleavage stage of in vitro fertilized embryos and cultured up to blastocyst. Also, porcine EG cells were labeled with DiO (Invitrogen, Carlsbad, CA) on the cell membrane or transfected with green fluorescent protein gene to observe whether the EG cells injected in the host embryo would incorporate into the inner cell mass (ICM) or trophectoderm (TE). Chimeric embryos were produced and allowed to develop into blastocysts to investigate the injected EG cells would come to lie in ICM and/or TE of the blastocyst, by scoring their position. In result, developmental rate was similar in all treatments. In all treatments, EG cells were mainly allocated in both ICM and TE of the chimeric blastocysts. These results suggest that examining the allocation pattern of injected EG cells, maintained pluripotency in vitro, could provide clues of differentiation process in vivo. Furthermore, to enhance the allocation of EG cells into the embryonic lineage, it would be required to optimize the culture condition for EG cells as well as embryos. Further experiment are needed to determine whether the injected EG cells could maintain their properties throughout the environment in the embryonic development in vitro. Table 1. Distribution of the porcine EG cells microinjected into cleavage-stage embryos

Stress-survival responses of a carbon-starvedp-nitrophenol-mineralizingMoraxellastrain in river water

2005 ◽  
Vol 51 (3) ◽  
pp. 223-229 ◽  
Author(s):  
Michael Moore ◽  
Jack Trevors ◽  
Hung Lee ◽  
Kam Tin Leung
Keyword(s):  
Green Fluorescent Protein ◽  
River Water ◽  
Water Samples ◽  
Protein Gene ◽  
Fluorescent Protein ◽  
Carbon Starvation ◽  
Green Fluorescent Protein Gene ◽  
River Water Samples ◽  
Stress Survival ◽  
Green Fluorescent

The effect of carbon starvation on the stress-resistant responses of a p-nitrophenol-mineralizing Moraxella strain was examined in both buffer and river water samples. The Moraxella strain showed optimal stress-resistant responses in a minimal salt buffer when carbon-starved for 1–2 d. In the buffer system, the 1- and 2-day carbon-starved Moraxella cultures survived about 150-, 200-, and 100-fold better than the non-starved cultures when exposed to 43.5 °C, 2.7 mol/L NaCl, and 500 µmol/L H2O2for 4 h, respectively. A green fluorescent protein gene- (gfp) labelled derivative of the Moraxella strain was used to examine the stress-resistant responses of the bacterium in natural river water microcosms. The carbon-starved gfp-labelled Moraxella strain also showed stress-resistant responses against heat, osmotic, and oxidative stresses in the river water samples. Despite the stress-tolerant capability of the carbon-starved gfp-labelled Moraxella cells, they did not exhibit any survival advantage over their non-starved counterparts when inoculated into river water microcosms and incubated at 10 and 22 °C for 14 d.Key words: carbon starvation, stress-survival responses, Moraxella, p-nitrophenol, green fluorescent protein gene.

Secretion of foreign proteins mediated by chicken lysozyme gene regulatory sequences

2002 ◽  
Vol 80 (6) ◽  
pp. 777-788 ◽  
Author(s):  
Gregory R Lampard ◽  
Ann M. Verrinder Gibbins
Keyword(s):  
Fluorescent Protein ◽  
Regulatory Sequences ◽  
Heterologous Proteins ◽  
Transgenic Chicken ◽  
Reporter Protein ◽  
Lysozyme Gene ◽  
Foreign Proteins ◽  
Gene Regulatory ◽  
Chicken Lysozyme

Exploitation of the insulating properties of the complete chicken lysozyme gene domain may facilitate the production of transgenic chicken bioreactors with the capacity to deposit valuable proteins in the egg white. Chimeric genes consisting of the chicken lysozyme gene regulatory sequences and sequences encoding foreign proteins could be inserted randomly into the chicken genome and retain appropriate expression levels. The research reported here established that chicken lysozyme gene regulatory sequences can be used to direct the production and secretion of green fluorescent protein (used as a reporter protein) in transiently transfected chicken blastodermal cells. Attempts to verify these findings in transgenic hens are currently in progress. To provide a rapid means of generating constructs encoding other foreign proteins under the control of lysozyme gene regulatory sequences that can facilitate the secretion of heterologous proteins in vivo, a generic lysozyme gene regulatory scaffold was created using a poxvirus-mediated gene targeting system.Key words: chicken lysozyme gene, secretion, homologous recombination.

Sign in / Sign up

Export Citation Format

Share Document