A Study of the Mitotically Active Germ Cells of the Gonads of X-Irradiated Chick Embryos

1968 ◽  
Vol 33 (2) ◽  
pp. 337
Author(s):  
S. M. Udgaonkar ◽  
B. K. Batra
Keyword(s):  
Germ Cells ◽  
Chick Embryos

Simple method for isolation of primordial germ cells from chick embryos

1992 ◽  
Vol 16 (9) ◽  
pp. 853-857 ◽  
Author(s):  
I CHANG ◽  
A TAJIMA ◽  
Y YASUDA ◽  
T CHIKAMUNE ◽  
T OHNO
Keyword(s):  
Germ Cells ◽  
Primordial Germ Cells ◽  
Chick Embryos ◽  
Simple Method

200 Optimization of Individual Stages of Chicken Transgenesis to Increase Efficiency

2018 ◽  
Vol 30 (1) ◽  
pp. 240
Author(s):  
E. K. Tomgorova ◽  
E. N. Antonova ◽  
N. A. Volkova ◽  
P. Y. Volchkov ◽  
N. A. Zinovieva
Keyword(s):  
Cell Population ◽  
Germ Cells ◽  
Cell Sorting ◽  
Optimal Dose ◽  
Cell Types ◽  
Dorsal Aorta ◽  
Chick Embryos ◽  
Embryonic Cells ◽  
Sex Cells ◽  
Adhesive Capacity

Primordial germ cells (PGC) are the precursors of male and female progenitor cells. The cells are considered a valuable genetic material for the production of transgenic poultry. This technology includes isolation of the PGC from chick donor embryos, transformation of the cells, and injection into the dorsal aorta of recipient embryos. After injection, the PGC are involved in the process of embryo development and differentiate into male or female sex cells. The aim of the research was to optimize the individual stages of this technology to increase the efficiency of transgenesis. The PGC were extracted from embryo gonads at stage 26 to 27 (H&H) using the trypsinization process. The trypsin concentration and incubation time were determined experimentally. Treatment of chick embryos with a 0.05% trypsin solution for 5 min was optimal for obtaining culture of embryonic cells. Separation of the PGC from other types of embryonic cells was based on a differential adhesive capacity. The maximum homogeneity of the cell population for further cultivation was established by transfer (twice) of the supernatant containing unattached cells after 1 h of cultivation in a new culture dish. The cell population is represented mainly by the PGC (81 ± 4%). Additional purification of the PGC from other cell types using magnetic-activated cell sorting (MACS) increased the proportion of these cells up to 93 ± 2%. The lentiviral transduction (pHAGE vector, ZsGreen under CMV promotor) was used to transform the resulting culture of the PGC. The efficiency of infection of PGC with lentiviral particles (TU/mL = 2.5 × 108) was 70 ± 3%. The transformed cells were injected into the dorsal aorta of recipient embryos on Day 2.5 (n = 80). Before injecting donor PGC, recipient embryos were treated with busulfan to remove the endogenous PGC. The optimal dose of busulfan was selected experimentally. A series of experiments introducing busulfan in concentrations from 50 to 250 μg into chick embryos at 24 h of incubation showed that the optimal dose was 100 μg/embryo. The efficacy of colonization of gonads with donor PGC was assessed on Day-10 embryos (n = 32) and 4-week-old hatched chickens (n = 12). Cells from gonads were studied using fluorescence microscopy, fluorescence-activated cell sorting (FACS) and qPCR. The presence of fluorescent cells in the gonads of recipients was established in both embryos and hatched chickens. The relative number of the recombinant DNA copies and the relative level of expression were confirmed by qPCR. The FACS analysis of sex cells isolated from gonads of recipients showed that the percentage of transformed germ cells reached 55.8% in females (n = 5) and 31.9% in males (n = 7). Thus, the effectiveness of poultry transgenesis can be enhanced by preparation of donor PGC for injection into embryo recipients and elimination of endogenous PGC in recipients. Both the purification of PGC from other cell types based on adhesive capacity as well as treatment of embryo recipients at 24 h incubation with busulfan (100 μg/embryo) increased the effectiveness of transgenesis. Study supported by the RSF within project No. 16-16-10059.

scholarly journals Genetic Factors Affect the Number of Circulating Primordial Germ Cells in Early Chick Embryos.

2003 ◽  
Vol 40 (2) ◽  
pp. 101-113 ◽  
Author(s):  
Dong-Feng Zhao ◽  
Hiroaki Yamashita ◽  
Masaharu Matsuzaki ◽  
Toshinori Takano ◽  
Shin-Ichi Abe ◽  
...  
Keyword(s):  
Germ Cells ◽  
Genetic Factors ◽  
Primordial Germ Cells ◽  
Chick Embryos

Analyse expérimentale de la régulation du nombre des cellules germinales après déficience gonadique chez le poulet

Development ◽  
1974 ◽  
Vol 32 (3) ◽  
pp. 619-635
Author(s):  
Eliane Didier ◽  
Nöel Fargeix ◽  
Yves Bergeaud
Keyword(s):  
Experimental Study ◽  
Germ Cells ◽  
Surgical Excision ◽  
Chick Embryos ◽  
Cellules Germinales ◽  
Control Chick ◽  
The Asymmetry

Experimental study of the regulation of the number of germ cells following gonadial deficiency in the chick Colonization of the genital ridges by germ cells was quantitatively studied in control chick embryos killed at stages 25–29, and in embryos in which a surgical excision of the gonad presumptive area was made previously on the second day. In operated embryos which show a more or less perfect agenesis of one gonad, the number of germ cells counted in genital ridges is lower than the number of germ cells estimated in the same stages of control embryos. The deficit is greater for left gonadic agenesis. The decrease in the total number of germ cells is essentially due to a reduction in the cells colonizing the deficient gonad. There is no excess of germ cells observed in the control gonad. Accordingly, a right side operation strengthens the asymmetry of germ cells distribution, whereas a left side one reduces it. Thus, in birds the regulation of the number of germ cells and the quantitative control of colonisation of the gonads is at the gonad level.

The role of the cell surface in the migration of primordial germ cells in early chick embryos: effects of concanavalin A

Development ◽  
1978 ◽  
Vol 46 (1) ◽  
pp. 5-20
Author(s):  
H. Lee ◽  
N. Karasanyi ◽  
R. G. Nagele
Keyword(s):  
Cell Surface ◽  
Germ Cells ◽  
Concanavalin A ◽  
Primordial Germ Cells ◽  
Sublethal Dose ◽  
Chick Embryos ◽  
Con A ◽  
And Migration ◽  
Coat Material

Effects of concanavalin A (Con A) on the morphology and migration of primordial germ cells (PGCs) in stage-6 to -12 chick embryos were investigated. Con A, at a sublethal dose (10µg/ml), inhibited migration of PGCs from the germinal crescent area to other parts of the embryo. Affected PGCs were more rounded without the usual cytoplasmic extensions, but the integrity of other structures was unaffected. Nearly identical results were obtained with another lectin, wheat germ agglutinin (10µg/ml). Histochemistry using Con A-horseradish peroxidase revealed that PGCs in control embryos had a thin, rather uniform layer of extracellular coat material (ECM). Con A appeared to alter the distribution of ECM on PGCs, i.e. some parts of the cell surface were devoid of any detectable ECM, while others had small, scattered patches of ECM. Con A effects were alleviated by α-methyl-d-mannoside. Overall results of the present study indicated that the observed inhibition of PGC migration in early chick embryos is a consequence of Con A-induced alterations of cell surface properties.

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