Skip to main content

Advertisement

SpringerLink
Log in

Efficient combined FISH and PRINS technique for detection of DAZ microdeletion in human sperm

  • Technological Innovations
  • Published:
Journal of Assisted Reproduction and Genetics Aims and scope Submit manuscript

Abstract

Intracytoplasmic sperm injection (ICSI) now offers an effective therapeutic option for men with male infertility and is believed to allow transmission of genetically determined infertility to the male offspring. Transmission of DAZ (Deleted in Azoospermia) microdeletion is one of the major concerns for oligo and severe oligozoospermia patients. Screening of the Y chromosome microdeletion in the diagnostic work-up of infertile men is mainly done using polymerase chain reaction (PCR) on blood leukocytes. However, there are evidences showing that presence of DAZ in somatic cells might not be indicative of its presence in germ cell lineage. In this report we are going to describe a combined Primed in situ labeling (PRINS) and fluorescence in situ hybridization (FISH) technique to show the localization of DAZ gene as well as Y chromosome centromere on sperm nuclei. PRINS is a combination of FISH and in situ polymerization provides another approach for in situ chromosomal detection. In the present study the PRINS primers specific for DAZ genes and traditional direct labeled centromere FISH probes for Y and X chromosomes were used in order to simultaneously detect DAZ genes and sex chromosome aneuploidy in sperm samples.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

References

  1. Krausz C, Degl’Innocenti S. Y chromosome and male infertility: update, 2006. Front Biosci. 2006;11:3049–61.

    Article  PubMed  CAS  Google Scholar 

  2. de Kretser DM. Male infertility. Lancet. 1997;349:787–90.

    Article  PubMed  Google Scholar 

  3. Krausz C, Murci LQ, McElreavey K. Prognostic value of Y chromosome microdeletion analysis. Hum Reprod. 2000;157:1431–4.

    Article  Google Scholar 

  4. Vogt PH. Human chromosome deletions in Yq11, AZF candidate genes and male infertility, history and update. Mol Hum Reprod. 1998;4:739–44.

    Article  PubMed  CAS  Google Scholar 

  5. Tiepolo L, Zuffardi O. Localization of factors controlling spermatogenesis in the nonfluorescent portion of the human Y chromosome long arm. Hum Genet. 1976;34:119–24.

    Article  PubMed  CAS  Google Scholar 

  6. Ma K, Inglis JD, Sharkey A, Bickmore WA, Hill RE, Prosser EJ, Speed RM, Thomson EJ, Jobling M, Taylor K, et al. A Y chromosome gene family with RNA-binding protein homology: candidates for the azoospermia factor AZF controlling human spermatogenesis. Cell. 1993;75:1287–95.

    Article  PubMed  CAS  Google Scholar 

  7. Reijo R, Lee TY, Salo P, Alagappan R, Brown LG, Rosenberg M, Rozen S, Jaffe T, Straus D, Hovatta O, et al. Diverse spermatogenic defects in humans caused by Y chromosome deletions encompassing a novel RNA-binding protein gene. Nat Genet. 1995;10:383–93.

    Article  PubMed  CAS  Google Scholar 

  8. Vogt PH, Edelmann A, Kirsch S, Henegariu O, Hirschmann P, Kiesewetter F, Kohn FM, Schill WB, Farah S, Ramos C, Hartmann M, Hartschuh W, Meschede D, Behre HM, Castel A, Nieschlag E, Weidner W, Grone HJ, Jung A, Engel W, Haidl G. Human Y chromosome azoospermia factors (AZF) mapped to different subregions in Yq11. Hum Mol Genet. 1996;5:933–43.

    Article  PubMed  CAS  Google Scholar 

  9. Krausz C, McElreavey K. Y chromosome and male infertility. Frontiers in Bioscience. 1999;4:1–8.

    Article  Google Scholar 

  10. Ferlin A, Arredi B, Speltra E, Cazzadore C, Selice R, et al. Molecular and clinical characterization of Y chromosome microdeletions in infertile men: a 10-year experience in Italy. J Clin Endocrinol Metabol. 2007;92:762–70.

    Article  CAS  Google Scholar 

  11. Navarro-Costa P, Gonçalves J, Plancha CE. The AZFc region of the Y chromosome: at the crossroads between genetic diversity and male infertility. Hum Reprod Update. 2010;16:525–42.

    Article  PubMed  CAS  Google Scholar 

  12. Vogt PH, Fernandes S: Polymorphic DAZ gene family in polymorphic structure of AZFc locus: Artwork or functional for human spermatogenesis? Acta Pathologica, Microbiologica et Immunologica Scandinavica 2003; 111 (online version).

  13. Meschede D, Lemcke B, Exeler JR, et al. Chromosome abnormalities in 447 couples undergoing intracytoplasmic sperm injection—prevalence, types, sex distribution and reproductive relevance. Hum Reprod. 1998;13:576–82.

    Article  PubMed  CAS  Google Scholar 

  14. Kostiner DR, Turek PJ, Reijo RA. Male infertility: analysis of the markers and genes on the human Y chromosome. Hum Reprod. 1998;13:3032–8.

    Article  PubMed  CAS  Google Scholar 

  15. Krausz C, Bussani-Mastellone C, Granchi S, McElreavey K, Scarselli G, Forti G. Screening for microdeletions of Y chromosome genes in patients undergoing intracytoplasmic sperm injection. Hum Reprod. 1999;14:1717–21.

    Article  PubMed  CAS  Google Scholar 

  16. Sharlip ID, Jarow JP, Belker AM, Lipshultz LI, Sigman M, Thomas AJ, Schlegel PN, Howards SS, Nehra A, Damewood MD, et al. Best practice policies for male infertility. Fertil Steril. 2002;5:873–82.

    Article  Google Scholar 

  17. Simoni S, Kamischke A, Nieschlag E. Significance of the molecular diagnosis of Y chromosomal microdeletions in the diagnostic workup of male infertility. Hum Reprod. 1998;13:1764–8.

    Article  PubMed  CAS  Google Scholar 

  18. Simoni M, Gromoll J, Dworniczak B, Rolf C, Abshagen K, et al. Screening for deletions of the Y chromosome involving the DAZ (Deleted in AZoospermia) gene in azoospermia and severe oligozoospermia. Fertil Steril. 2010;67:1753–6.

    Google Scholar 

  19. Zini A, Libman J. Sperm DNA damage: clinical significance in the era of assisted reproduction. Can Med Assoc J. 2006;175:495–500.

    Article  Google Scholar 

  20. Nili HA, Mozdarani H, Aleyasin A. Correlation of sperm DNA damage with protamine deficiency in Iranian subfertile men. Reprod Biomed Online. 2009;18:479–85.

    Article  PubMed  Google Scholar 

  21. Aitken RJ, Ryan AL, Curry BJ, Baker MA. Multiple forms of redox activity in populations of human spermatozoa. Mol Hum Reprod. 2003;9:645–61.

    Article  PubMed  CAS  Google Scholar 

  22. Deepali P, Snjay P, Jyoti S, Sebastian PC, Sher A. Genomic instability of the DYZ1 repeat in patient with Y chromosome anomalies and males exposed to natural background radiation. DNA Res. 2006;13:103–9.

    Article  Google Scholar 

  23. Snjay P, Jyoti S, Sebastian PC, Ahamd J, Sher A. Tandem duplication and copy number polymorphism of the SRY gene in patients with sex chromosome anomalies and males exposed to natural background radiation. Mol Hum Reprod. 2006;12:113–21.

    Article  Google Scholar 

  24. Snjay P, Jyoti S, Sebastian PC, Sher A. AZFc somatic microdeletions and copy number polymorohism of the DAZ genes in human males exposed to natural background radiation. Hum Genet. 2007;121:337–46.

    Article  Google Scholar 

  25. Arruda JT, Silva DM, Silva CC, Moura KKVO, da-Cruz AD. Homologous recombination between HERVs causes duplications in the AZFa region of men accidentally exposed to cesium-137 in Goiânia. Genet Mol Res. 2008;7:1063–9.

    Article  PubMed  CAS  Google Scholar 

  26. Arruda JT. Occurrence of mutations in loci linked to Y chromosome in the offspring born to individuals exposed to ionizing radiation. Genet Mol Res. 2009;8:938.

    Article  CAS  Google Scholar 

  27. Moghbeli-Nejad S, Mozdarani H, Behmanesh M, Rezaiean Z, Fallahi P. Genome instability in AZFc region on Y chromosome in leukocytes of fertile and infertile individuals following exposure to gamma radiation. J Assist Reprod Genet. 2012;29:53–61.

    Article  PubMed  Google Scholar 

  28. Kent-First MG, Kol S, Muallem A, Ofir R, Manor D, Blazer S, First N, Itskovitz-Eldor J. The incidence and possible relevance of Y-linked microdeletions in babies born after intracytoplasmic sperm injection and their infertile fathers. Mol Hum Reprod. 1996;2:943–50.

    Article  PubMed  CAS  Google Scholar 

  29. Dada R, Kumar R, Shamsi MB, Kumar R, Kucheria K, Sharma RK, Gupta SK, Gupta NP. Higher frequency of Yq microdeletions in sperm DNA as compared to DNA isolated from blood. Asian J Androl. 2007;9:720–2.

    Article  PubMed  CAS  Google Scholar 

  30. Sakthivel PJ, Swaminathan M. Y chromosome microdeletions in sperm DNA of infertile patients from Tamil Nadu, south India. Indian J Urol. 2008;24:480–5.

    Article  PubMed  Google Scholar 

  31. Le Bourhis C, Siffroi JP, McElreavey K, Dadoune JP. Y chromosome microdeletions and germinal mosaicism in infertile males. Mol Hum Reprod. 2000;6:688–93.

    Article  PubMed  Google Scholar 

  32. De Vries JW, Repping S, Oates R, Carson R, Leschot NJ, Van Der Veen F. Absence of deleted in azoospermia (DAZ) genes in spermatozoa of infertile men with somatic DAZ deletions. Fertil Steril. 2001;75:476–9.

    Article  PubMed  Google Scholar 

  33. Gao J-L, Nie Y, Ding X-P. Primed in situ labeling for detecting single-copy genes. Genet Mol Res. 2011;10:1884–90.

    Article  PubMed  CAS  Google Scholar 

  34. Kadandale JS, Wachtel SS, Tunca Y, Martens PR, et al. Deletion of RBM and DAZ in azoospermia: evaluation by PRINS. Am J Med Genet. 2002;107:105–8.

    Article  PubMed  Google Scholar 

  35. Tharapel AT, Wachtel SS. PRINS for mapping single-copy genes. Methods Mol Biol. 2006;338:59–67.

    PubMed  CAS  Google Scholar 

  36. Pellestor F, Girardet A, Lefort G, Andreo B, et al. PRINS as a method for rapid chromosomal labeling on human spermatozoa. Mol Reprod Dev. 1995;40:333–7.

    Article  PubMed  CAS  Google Scholar 

  37. Koch JE, Kolvraa S, Petersen KB, Gregersen N, Bolund L. Oligonucleotide-priming methods for the chromosome-specific labeling of alpha satellite DNA in situ. Chromosoma. 1989;98:259–65.

    Article  PubMed  CAS  Google Scholar 

  38. Pellestor F. What PRINS can do for you. Medical Science. 1998;14:935–8.

    Google Scholar 

  39. World Health Organization: WHO laboratory manual for the Examination and processing of human semen. Fifth edition. WHO Press, 2010.

  40. Aitken RJ, Clarkson JS. Significance of reactive oxygen species and antioxidants in defining the efficacy of sperm preparation techniques. J Androl. 1988;9:367–76.

    PubMed  CAS  Google Scholar 

  41. Pellestor F, Girardet A, Coignet L, Andréo B, Charlieu J-P. Assessment of aneuploidy for chromosomes 8, 9, 13, 16 and 21 in human sperm by using primed in situ labeling technique. Am J Hum Genet. 1996;58:797–802.

    PubMed  CAS  Google Scholar 

  42. Pellestor F, Paulasova P, Andréo B, Lefort G, Hamamah S. Multicolor PRINS and multicolor PNA. Cytogenet Genom Res. 2006;114:263–9.

    Article  CAS  Google Scholar 

  43. Yan J, Bronsard M, Drouin R. Creating a new color by omission of 3 end blocking step for simultaneous detection of different chromosomes in multi-PRINS technique. Chromosoma. 2001;109:565–70.

    Article  PubMed  CAS  Google Scholar 

  44. Writzl K, Zorn B, Peterlin B. Copy number of DAZ genes in infertile men. Fertil Steril. 2005;84:1522–5.

    Article  PubMed  CAS  Google Scholar 

  45. Noordam MJ, Westerveld GH, Hovingh SE, van Daalen SK, Korver CM, van der Veen F, van Pelt AM, Repping S. Gene copy number reduction in the azoospermia factor c (AZFc) region and its effect on total motile sperm count. Hum Mol Genet. 2011;20:2457–63.

    Article  PubMed  CAS  Google Scholar 

  46. Barrat CL, Aitken RJ, Björndahl L, Carrell DT, de Boer P, et al. Sperm DNA: organization, protection and vulnerability: from basic science to clinical applications–a position report. Hum Reprod. 2010;25:824–38.

    Article  Google Scholar 

Download references

Acknowledgements

This research was supported by the Research Department of the Faculty of Medical Sciences, Tarbiat Modares University. The authors would like to express their thanks to Mrs Z. Rezaeian for arrangements for sample collection.

Conflict of Interest statement

The authors declare that there are no conflicts of interest.

Author information

Authors and Affiliations

  1. Department of Medical Genetics, Faculty of Medical Sciences , Tarbiat Modares University, P.O. Box: 14115-111, Tehran, Iran

    Hossein Mozdarani & Pegah Ghoraeian

Authors
  1. Hossein Mozdarani
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Pegah Ghoraeian
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Hossein Mozdarani.

Additional information

Capsule

Using combined techniques of PRINS and FISH, both DAZ localization and Y chromosome centromere on human sperm can be visualized easily on single cells.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Esm 1

(PDF 198 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Mozdarani, H., Ghoraeian, P. Efficient combined FISH and PRINS technique for detection of DAZ microdeletion in human sperm. J Assist Reprod Genet 29, 979–984 (2012). https://doi.org/10.1007/s10815-012-9805-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10815-012-9805-z

Keywords

Search

Navigation