Poultry genetic resource conservation using primordial germ cells

Nakamura, Yatsuka

doi:10.1262/jrd.2016-052

Cited by 31 publications

(19 citation statements)

References 81 publications

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“…Since in chicken the males are the homogametic sex, it needs a multigenerational breeding program to reconstitute a breed only with the use of sperm 45 . The reconstitution of a breed using PGCs is based on the production of germline chimeras which can be mated with other chimeras, living individuals or artificially inseminated with stored sperm 46 , which will drastically shorten the time to reconstitute a breed or transfer a rare allele into a recipient chicken line.…”

Section: Discussionmentioning

confidence: 99%

Cultivation and characterization of primordial germ cells from blue layer hybrids (Araucana crossbreeds) and generation of germline chimeric chickens

Altgilbers

Klein

Dierks

et al. 2021

Sci Rep

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The chicken (Gallus gallus) is one of the most common and widespread domestic species, with an estimated total population of 25 billion birds worldwide. The vast majority of chickens in agriculture originate from hybrid breeding programs and is concentrated on few commercially used high performance lines, whereas numerous local and indigenous breeds are at risk to become extinct. To preserve the genomic resources of rare and endangered chicken breeds innovative methods are necessary. Here, we established a solid workflow for the derivation and biobanking of chicken primordial germ cells (PGCs) from blue layer hybrids. To achieve this, embryos of a cross of heterozygous blue egg layers were sampled to obtain blood derived and gonadal male as well as female PGCs of different genotypes (homozygous, heterozygous and nullizygous blue-allele bearing). The total efficiency of established PGC lines was 45% (47/104) within an average of 49 days until they reached sufficient numbers of cells for cryopreservation. The stem-cell character of the cultivated PGCs was confirmed by SSEA-1 immunostaining, and RT-PCR amplification of the pluripotency- and PGC-specific genes cPOUV, cNANOG, cDAZL and CVH. The Sleeping Beauty transposon system allowed to generate a stable integration of a Venus fluorophore reporter into the chicken genome. Finally, we demonstrated that, after re-transfer into chicken embryos, Venus-positive PGCs migrated and colonized the forming gonads. Semen samples of 13 raised cell chimeric roosters were analyzed by flow cytometry for the efficiency of germline colonization by the transferred PGCs carrying the Venus reporter and their proper differentiation into vital spermatids. Thus, we provide a proof-of-concept study for the potential use of PGCs for the cryobanking of rare breeds or rare alleles.

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Section: Discussionmentioning

confidence: 99%

Cultivation and characterization of primordial germ cells from blue layer hybrids (Araucana crossbreeds) and generation of germline chimeric chickens

Altgilbers

Klein

Dierks

et al. 2021

Sci Rep

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“…However, it is impossible to cryopreserve avian ova. PGCs are an ideal alternative to preserve the germplasm of both sexes (Nakamura, 2016 ). However, the isolation and in vitro culture of waterfowl PGCs have been rarely described, in contrast with the many studies of chicken PGCs, and there are only a few reports concerning germ cells of waterfowls.…”

Section: Introductionmentioning

confidence: 99%

In vitro culture and characterization of duck primordial germ cells

et al. 2019

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This study aimed to isolate, culture, and characterize duck primordial germ cells (PGCs) and to compare these cells with chicken PGCs. We first cultured Muscovy duck (Cairina moschata) circulating PGCs and gonadal PGCs (gPGCs) in the modified serum-containing medium used to amplify chicken PGCs. gPGCs were found to proliferate better in serum-free chemically defined medium than in serum-containing medium. Thereafter, gPGCs were similarly isolated from 2 other duck breeds, the Pekin duck (Anas platyrhynchos) and the hybrid mule duck (C. moschata × A. platyrhynchos), and amplified for a limited period of time in the chemically defined culture condition, but sufficiently to be characterized and transplanted. Cultured gPGCs of all 3 duck breeds were characterized by Periodic acid-Schiff staining, immunocytochemical staining, and expression analysis of germline-specific and pluripotency genes. Cultured duck gPGCs colonized the gonads after being genetically labeled and injected into recipient embryos. Taken together, these results demonstrate that duck PGCs retain their germline characteristics after being isolated, expanded in vitro, and genetically modified. Further studies are required to establish the optimal conditions for long-term culture of duck PGCs, which may involve supplementing the culture medium with other growth factors or compounds.

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“…The more feasible and available method for management of reproductive cells in bird populations is, therefore, semen cryopreservation (Blesbois et al., 2007). Other methods including primordial germ cells and gonadic tissues technologies are still in progress but may be seen as methods that are complementary to semen cryopreservation (Liptoi et al., 2013; Nakamura, 2016; Sztan et al., 2017; Thélie et al., 2018). Different freezing procedures have been developed to cryopreserve avian semen (Blesbois et al., 2007; Santiago-Moreno et al., 2012; Rakha et al., 2016, 2017a; 2018a,b,c; Thananurak et al., 2016; Chuaychu-noo et al., 2017; Thananurak et al., 2017).…”

Section: Introductionmentioning

confidence: 99%

Sucrose increases the quality and fertilizing ability of cryopreserved chicken sperms in contrast to raffinose

Thananurak¹,

Chuaychu-noo²,

Thélie³

et al. 2019

Poultry Science

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Chicken semen conservation is an important tool for programs of genetic diversity management and of endangered breeds’ conservation. However, the method still needs to be improved in order to be applied in a wide variety of environments and breeds. Our objective was to compare the effects of 2 external cryoprotectants saccharides (sucrose and raffinose) on the sperm freezability of a Thai local breed, Pradu Hang Dum, in which semen was frozen with a simple freezing method using nitrogen vapors and dimethyl formamide (DMF). Thirty-six males were selected on their motility vigor score for the experiments. In a first experiment, a large range of sucrose and raffinose doses were tested. Semen quality was evaluated after incubation at 5°C or after cryopreservation in straws in the saline Blumberger Hahnen Sperma Verdünner diluent + DMF (6% v/v) with or without sucrose/raffinose. The best targeted doses of sucrose and raffinose were then kept for experiment 2 that was focused on cryopreserved semen. In this experiment, semen quality was measured on frozen-thawed sperm: different objective motility data evaluated by computer-assisted sperm analysis (CASA), membrane integrity, acrosome integrity, mitochondria function evaluated using flow cytometry, lipid peroxide production assessed by the thiobarbituric acid test. Fertility obtained with frozen-thawed semen supplemented or not with sucrose or raffinose was also evaluated after artificial insemination of laying hens. The presence of sucrose at the osmotically inactive dose 1 mmol significantly increased the vigor motility, membrane integrity, acrosome integrity, and mitochondrial functions of frozen-thawed sperm ( P < 0.05), and showed the highest levels of fertility after sperm cryopreservation (91% vs. control 86%, P < 0.001). Raffinose showed negative effects on in vitro semen quality from 1 to 100 mmol. Fertility was also negatively ( P < 0.001) affected by raffinose (fertility rate 66 to 70%). We thus showed in the present study the high success of a simple chicken sperm cryopreservation method with an external cryoprotectant easily available and cheap, the sucrose, used at an osmotically inactive low concentration.

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Poultry genetic resource conservation using primordial germ cells

Cited by 31 publications

References 81 publications

Cultivation and characterization of primordial germ cells from blue layer hybrids (Araucana crossbreeds) and generation of germline chimeric chickens

Cultivation and characterization of primordial germ cells from blue layer hybrids (Araucana crossbreeds) and generation of germline chimeric chickens

In vitro culture and characterization of duck primordial germ cells

Sucrose increases the quality and fertilizing ability of cryopreserved chicken sperms in contrast to raffinose

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