• Title/Summary/Keyword: germ line

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Essential Role of brc-2 in Chromosome Integrity of Germ Cells in C. elegans

  • Ko, Eunkyong;Lee, Junho;Lee, Hyunsook
    • Molecules and Cells
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    • v.26 no.6
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    • pp.590-594
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    • 2008
  • brc-2, an ortholog of BRCA2 in Caenorhabditis elegans, is essential in the maintenance of genetic integrity. In C. elegans, cellular location correlates with meiotic progression, and transgene-induced cosuppression is observed in the germ line but not in somatic cells. We used these unique features to dissect the role of brc-2 in the germ line from that in somatic cells. In situ hybridization of wild type animals revealed that brc-2 gene expression was higher in oocytes than in other germline cells, and was barely detectable in mitotic cells. In contrast, germ cells containing multicopies of the brc-2 transgene showed no significant in situ hybridization signal at any oogenesis stage, confirming that brc-2 expression was functionally cosuppressed in the transgenic germ line. RAD-51 foci formation in response to DNA damage was abrogated in brc-2-cosuppressed germ cells, whereas wild-type germ cells showed strong RAD-51 foci formation. These germ cells exhibited massive chromosome fragmentation and decompaction instead of six bivalent chromosomes in diakinesis. Accordingly, lethality was observed after the early stage of germline development. These results suggest that brc-2 plays essential roles in chromosome integrity in early prophase, and therefore is crucial in meiotic progression and embryonic survival.

A Mutation of cdc-25.1 Causes Defects in Germ Cells But Not in Somatic Tissues in C. elegans

  • Kim, Jiyoung;Lee, Ah-Reum;Kawasaki, Ichiro;Strome, Susan;Shim, Yhong-Hee
    • Molecules and Cells
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    • v.28 no.1
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    • pp.43-48
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    • 2009
  • By screening C. elegans mutants for severe defects in germline proliferation, we isolated a new loss-of-function allele of cdc-25.1, bn115. bn115 and another previously identified loss-of-function allele nr2036 do not exhibit noticeable cell division defects in the somatic tissues but have reduced numbers of germ cells and are sterile, indicating that cdc-25.1 functions predominantly in the germ line during postembryonic development, and that cdc-25.1 activity is probably not required in somatic lineages during larval development. We analyzed cell division of germ cells and somatic tissues in bn115 homozygotes with germline-specific anti-PGL-1 immunofluorescence and GFP transgenes that express in intestinal cells, in distal tip cells, and in gonadal sheath cells, respectively. We also analyzed the expression pattern of cdc-25.1 with conventional and quantitative RT-PCR. In the presence of three other family members of cdc-25.1 in C. elegans, defects are observed only in the germ line but not in the somatic tissues in cdc-25.1 single mutants, and cdc-25.1 is expressed predominantly, if not exclusively, in the germ line during postembryonic stages. Our findings indicate that the function of cdc-25.1 is unique in the germ line but likely redundant with other members in the soma.

Transgenesis and Germ Cell Engineering in Domestic Animals

  • Lee, C.K.;Piedrahita, J.A.
    • Asian-Australasian Journal of Animal Sciences
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    • v.16 no.6
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    • pp.910-927
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    • 2003
  • Transgenesis is a very powerful tool not only to help understanding the basics of life science but also to improve the efficiency of animal production. Since the first transgenic mouse was born in 1980, rapid development and wide application of this technique have been made in laboratory animals as well as in domestic animals. Although pronuclear injection is the most widely used method and nuclear transfer using somatic cells broadens the choice of making transgenic domestic animals, the demand for precise manipulation of the genome leads to the utilization of gene targeting. To make this technique possible, a pluripotent embryonic cell line such as embryonic stem (ES) cell is required to carry genetic mutation to further generations. However, ES cell, well established in mice, is not available in domestic animals even though many attempt to establish the cell line. An alternate source of pluripotent cells is embryonic germ (EG) cells derived from primordial germ cells (PGCs). To make gene targeting feasible in this cell line, a better culture system would help to minimize the unnecessary loss of cells in vitro. In this review, general methods to produce transgenic domestic animals will be mentioned. Also, it will focus on germ cell engineering and methods to improve the establishment of pluripotent embryonic cell lines in domestic animals.

Methylenetetrahydrofolate Reductase Gene Germ-Line C677T and A1298C SNPs are Associated with Colorectal Cancer Risk in the Turkish Population

  • Ozen, Filiz;Sen, Metin;Ozdemir, Ozturk
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.18
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    • pp.7731-7735
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    • 2014
  • Colorectal cancer (CRC) is the third most common cause of death due to cancer in the worldwide and the incidence is also increasing in Turkey. Our present aim was to investigate any association between germ-line methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C polymorphisms and CRC risk in Turkey. A total of 86 CRC cases and 212 control individuals of the same ethnicity were included in the current study. Peripheral blood-DNA samples were used for genotyping by StripAssay technique, based on the reverse-hybridization principle and real-time PCR methods. Results were compared in Pearson Chi-square and multiple logistic regression models. The MTHFR 677TT (homozygous) genotype was found in 20.9% and the T allele frequency 4.2-fold increased in CRC when compared with the control group.The second SNP MTHFR 1298CC (homozygous) genotype was found in 14.0% and the C allele frequency 1.4-fold elevated in the CRC group. The current data suggest strong associations between both SNPs of germ-line MTHFR 677 C>T and 1298 A>C genotypes and CRC susceptibility in the Turkish population. Now the results need to be confirmed with a larger sample size.

Combined Germline Variations of Thrombophilic Genes Promote Genesis of Lung Cancer

  • Ozen, Filiz;Polat, Fikriye;Arslan, Sulhattin;Ozdemir, Ozturk
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.9
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    • pp.5449-5454
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    • 2013
  • Background: A large variety of familiar and non-familiar lung carcinomas (LC) are caused by long term exposure to chemical carcinogens that are present in tobacco smoke. We aimed to investigate the prevalence of 5 thrombophilic germ-line mutations in patients with lung carcinomas. Materials and Methods:A total of 52 LC patients and 212 healthy controls from same population were analyzed for FV Leiden, factor V H1299R (R2), PAI-1, MTHFR C677T, MTHFR A1298C, ACE I/D, and Apo E genes and compared. Results: Overall, heterozygous and/or homozygous point mutations in FV Leiden Apo E2, PAI-1 and MTHFR C677T genes were associated with LC in the current cohort. There was no meaningful association between LC and ACE I/D gene markers. Conclusions: The current results showed that LC is related to combined thrombophilic gene mutations and individuals with homozygosity of 4G in PAI-1 and MTHFR C677T genes and heterozygosity of FV Leiden, Apo E4 genes have a germ-line risk for LC tumorigenesis.

GENE TRANSFER BY MANIPULATION OF PRIMORDIAL GERM CELLS IN THE CHICKEN

  • Han, Jac Y.;Shoffner, R.N.;Guise, K.S.
    • Asian-Australasian Journal of Animal Sciences
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    • v.7 no.3
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    • pp.427-434
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    • 1994
  • The primordial germ cells (PGCs) were transfected in vitro and expressed the exogenous RSVLTR/${\beta}G2$ plasmid, suggesting thaI PGC is a possible vector for direct gene transfer into the germ line. Transfection efficiency of cell suspensions containing PGCs was 1.5% by liposome mediated DNA transfection. By microinjection of the transfected PGCs into the host germinal crescent, PGCs migrated via blood vessel to the future gonad and these transfected PGCs resulted in the RSVLTR/${\beta}G2$ expression in the gonad. The results from the seeding of PGCs on the chorioallantoic membrane were insufficient to test the hypothesis that PGCs can penetrate or invade the chorioallantoic membrane for transport via the circulatory system.

Potential Reproductive Toxicity Study of p53 Expressing Adenoviral Vector in Mice (아데노바이러스 유전자치료벡터의 생식독성 연구)

  • Rhee, Gyu-Seek;Kwack, Seung-Jun;Kim, Soon-Sun;Lee, Rhee-Da;Seok, Ji-Hyun;Chae, Soo-Young;Chung, Soo-Youn;Kim, Seung-Hee;Lee, Seung-Hoon;Park, Kui-Lea
    • Korean Journal of Microbiology
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    • v.43 no.3
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    • pp.151-158
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    • 2007
  • The possibility of inadvertent introduction of therapeutic gene expressing viral vectors has raised safety concerns about germ-line infection. Particularly, for indications such as prostate cancer and ovarian cancer, the proximity of the point of viral administration to organs of the reproductive system raises concerns regarding inadvertent germ-line transmission of genes carried by the virus vector. To evaluate the safety of in vivo adenovirus mediated gene transfer, we explored the biodistribution, persistance and potential germ-line transmission of p53-expressing adenovirus (Ad-CMV-p53). Both male and female Balb/c mice were injected with $1{\times}10^9$ PFU of Ad-CMV-p53. The PCR analysis showed that there were detectable vector sequences in liver, kidney, spleen, seminal vesicle, epididymis, prostate, ovary, and uterus. The RT-PCR analysis for detecting inserted gene, p53 showed that Ad-CMV-p53 viral RNA were present in spleen, prostate and ovary. Direct injected male and female mice of adenovirus vector into testis and ovary were mated and their of offspring were evaluated for germ-line transmission of the adenoviral vector. The PCR and RT-PCR analysis showed no evidence of germline transmission, although vector sequences were detected in DNA extracted from gonadal tissues. Real-time PCR result confirmed a significant decrease of adenovirus in gonad tissues 1 week after injection. We have also analysed the cell specific localization of viral DNA in gonad tissues by using in-situ PCR. Positive signals were detected in interstitial tissue but not in seminiferous tubule in sperm. In the case of ovary, adenovirus signal were localized to the stromal tissue, but no follicular signals were observed. Together, these data provide strong evidence that the risk of the Inadvertent germ-line transmission of vector sequences following intraperitoneal or direct injection into genito-urinary system of adenovirus is extremely low.

Preparation and Characterization of Wheat Germ Agglutinin-conjugated Praecoxin A (Wheat Germ Agglutinin-conjugated Praecoxin A의 제작 및 특성)

  • 김완수;김만석;김범수;이민원;이도익
    • YAKHAK HOEJI
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    • v.45 no.3
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    • pp.302-309
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    • 2001
  • Wheat germ agglutinin (WGA) pectin, which binds to human melanoma cell line, was conjugated with Praecoxin A using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide as a cross-linking agent. Physical mixture (PM) of WGA and Praecoxin A was also prepared by using a non-specific binding property of Praecoxin A to WGA. The WGA:Praecoxin A ratio in the conjugate and PM was approximately 1:18 and 1:20, respectively. The results of hemagglutination assay and enzyme-linked lectin assay indicated that the conjugate and PM maintained the lectin-like properties of the WGA. The binding ratio of conjugate was about 70% during 4-24 hr, but the most of Praecoxin A was released within 24 hr in the case of PM. These results lead to the conclusion that the conjugate is potentially useful for the formulation of injection that requires targeting for melanoma as well as sustained release at the site.

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