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Sexual Differentiation: Immunological Aspects
Published in Gérard Chaouat, The Immunology of the Fetus, 2020
Joyce A. Shelton, Erwin Goldberg
Page and co-workers61 have cloned a 230-kilobase segment of the human Y-chromosome that may contain all or part of the gene for TDF. The sequence of this segment appears to be highly conserved in mammals and birds. The structure of this portion of the DNA resembles that of frog transcription factor IIIA and, therefore, may be coding for a protein that binds nucleic acids and not for a cell surface protein. Presumably, this protein, which has been designated ZFY for “Y-bome zinc finger protein”, would not be accessible to antibody or cellular recognition. It, therefore, represents yet another candidate, quite separate from H-Y or SDM antigens, for the testis-inducing factor. Another report suggests that the ZFY protein may not be the primary inductive signal.62 The DNA probe for ZFY detects a homologous sequence in marsupials, but on an autosome rather than on the Y-chromosome. The Y-chromosome is necessary for male sexual differentiation in marsupials; consequently, the autosomal location of ZFY-coding sequences implies that, at least in marsupials, it is not a primary factor in male sex determination. A less direct involvement for ZFY, perhaps a regulatory function, has not been ruled out. The role of this protein in other species may need to be further evaluated.
Discovery and research
Published in Peter S. Harper, The Evolution of Medical Genetics, 2019
… there were three groups, myself, Jean Weissenbach and David Page, who were left with the tools and the inclination to try and clone the sex determining gene, and essentially we were all using the same approach, which was to construct a map across the Y chromosome using DNA from XX males and XY females to try and identify, classic positional cloning, the minimal region where the gene ought to be. And MIC2, which we had cloned and shown to be pseudo-autosomal and mapped it, and cloned the pseudoautosomal boundary, we knew that it was the closest marker distally to the gene, and so we started walking. And we'd identified a CPG island by using pulsed-field gel electrophoresis which we thought must be the next gene down. And it was the next gene down and it was a gene called ZFY and basically we had just isolated it when we read David Page's paper saying that he had cloned this gene and he had been walking from below it towards it, and that he presented evidence that it was the sex determining gene.
Prenatal Diagnosis
Published in Victor A. Bernstam, Pocket Guide to GENE LEVEL DIAGNOSTICS in Clinical Practice, 2019
Another PCR-based assay for fetal sexing identifies a single and discrete 530-bp fragment using two oligonucleotide primers specific for the ZFY gene cloned from a region of the human Y chromosome. While these procedures are definitely attractive for a number of clinical and forensic uses when sex determination is sought, their application to maternal blood, although possible, has not yet been reported.
Differentiation of spermatogonial stem cells by soft agar three-dimensional culture system
Published in Artificial Cells, Nanomedicine, and Biotechnology, 2019
Elham Mohammadzadeh, Tooba Mirzapour, Mohammad Reza Nowroozi, Hamid Nazarian, Abbas Piryaei, Fatemeh Alipour, Sayed Mostafa Modarres Mousavi, Marefat Ghaffari Novin
The obtained cells suspension (a mixture of spermatogonial cells and Sertoli cells) were filtered through a 70 µm nylon filter and cultured for three weeks at 37 °C, in the presence of 10% fetal bovine serum (FBS; Thermo Fisher scientific),1% Pen/Strep and 5% CO2, in a humidified atmosphere [16]. After 3 weeks, the cells were dissociated by (EDTA)–trypsin treatment (0.02% EDTA–0.1% trypsin) at 37° C for 5 min and the presence of spermatogonial stem cells in culture system was confirmed by immunocytochemistry of GFR-α1 and ITGα6 antibodies. Then, the cell suspension was cultured on SACS, gelatin-coated plates and control groups for another two weeks. In this study, the secretions of Sertoli cells were extracted from obstructive azoospermia (OA) patients and used as a conditioned medium for culturing the germ cells of non-obstructive azoospermia (NOA) patients in three groups. The number and diameter of colonies were evaluated on days 3, 7, 10 and 14 after culture using an inverted microscope (Olympus IX71, EXFO 120 Xenon-Hg excitation light). The expression of Oct4, Stra8, Scp3 and Acrosin genes was also evaluated before and after 2 weeks of culture by quantitatively real-time PCR. The Y chromosome microdeletions (including SRY, ZFY, sY84, sY86, sY127, sY134, sY254 and sY255) were investigated in NOA patients by multiplex PCR technique.
High frequency of de novo DAZ microdeletion in sperm nuclei of subfertile men: possible involvement of genome instability in idiopathic male infertility
Published in Human Fertility, 2018
Hossein Mozdarani, Pegah Ghoraeian, Sohail Mozdarani, Parvin Fallahi, Anahita Mohseni-Meybodi
From each man, 5 ml blood was collected in the heparinized tubes. Metaphase spreads were obtained from phytohemaglutinin (PHA, Gibco, BRL, Carlsbad, CA) stimulated cultured lymphocytes after 72 h and using standard techniques; cytogenetic analysis of all donors was done on G-banded chromosomes. The patients with 46 XY, normal karyotypes were selected for DAZ microdeletion analysis. Peripheral blood samples were collected in tubes containing anti-coagulant ethylenediaminetetraacetic acid (EDTA) from cytogenetically normal infertile patients. All tests were performed on genomic DNA that was extracted from peripheral blood leukocytes using a commercially available DNA-isolation kit for mammalian blood (Cinna Puregene, Tehran, Iran). For each participant, DAZ-specific STSs that spanned the subregions (sY254 and sY255) as well as STS for SRY and ZFY/ZFX as internal controls were used to amplify specific regions of the Y chromosome using PCR (Simoni, Bakker, & Krausz, 2004). The PCR amplification comprised a total volume of 25 μl, containing 200 ng of DNA; and standard PCR reaction mixture (all materials from Fermentas, Waltham, MA) and thermocycling. The PCR reaction products were, separated on 2–3% (w/v) agarose gels, and visualized by staining the gel with ethidium bromide. Negative and positive samples were judged based on a negative and positive control.
Non-invasive prenatal diagnosis of foetal gender through maternal circulation in first trimester of pregnancy
Published in Journal of Obstetrics and Gynaecology, 2019
Saeed Mahdavi, Fatemeh Karami, Saeed Sabbaghi
Foetal sex was correctly determined in total 147/147 pregnancies which have been successfully followed up until the birth and the remaining 45 cases have been excluded from this study due to failure in post-natal determination of neonatal gender. Co-amplification of two SRY and ZFY genes was detected in 103 male foetuses (Table 1).