{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T04:11:03Z","timestamp":1787026263837,"version":"build-2736575974"},"reference-count":78,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2021,6,14]],"date-time":"2021-06-14T00:00:00Z","timestamp":1623628800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Cell Dev. Biol."],"abstract":"<jats:p>\n                    Sperm quality is usually evaluated prior to artificial insemination in farm animals. In addition to conventional semen analysis, other biomarkers, such as mitochondrial activity, integrity and lipid disorder of plasma membrane, generation of reactive oxygen species (ROS) and sperm DNA integrity, have been found to be related to fertility rates in different species. While mounting evidence indicates that the Comet assay is a sensitive method for the detection of DNA breaks, complete sperm chromatin decondensation is required in order to properly analyze the presence of single- and double-strand DNA breaks. In this sense, a previous study showed that longer lysis treatment with proteinase K is needed to achieve complete chromatin decondensation. The current work sought to determine which specific lysis treatment leads to complete chromatin decondensation in pig sperm, as this is needed for the measurement of DNA damage in this species. With this purpose, incubation with a lysis solution containing proteinase K for 0, 30, and 180 min was added to the conventional protocol. The impact of the DNA damage induced by hydrogen peroxide (H\n                    <jats:sub>2<\/jats:sub>\n                    O\n                    <jats:sub>2<\/jats:sub>\n                    ; 0.01 and 0.1%) and DNAse I (1U and 4U) was also evaluated. Complete chromatin decondensation was only achieved when a long additional lysis treatment (180 min) was included. Furthermore, olive tail moment (OTM) and percentage of tail DNA (TD) indicated that a higher amount of DNA breaks was detected when hydrogen peroxide and DNAse I treatments were applied (\n                    <jats:italic>P<\/jats:italic>\n                    &amp;lt; 0.05). The comparison of treated and control sperm allowed defining the thresholds for OTM; these thresholds revealed that the percentage of sperm with fragmented DNA determined by the alkaline Comet does not depend on chromatin decondensation (\n                    <jats:italic>P<\/jats:italic>\n                    &amp;gt; 0.05). In conclusion, complete chromatin decondensation prior to alkaline and neutral Comet assays is needed to analyze DNA breaks in pig sperm.\n                  <\/jats:p>","DOI":"10.3389\/fcell.2021.675973","type":"journal-article","created":{"date-parts":[[2021,6,14]],"date-time":"2021-06-14T02:58:12Z","timestamp":1623639492000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":13,"title":["Complete Chromatin Decondensation of Pig Sperm Is Required to Analyze Sperm DNA Breaks With the Comet Assay"],"prefix":"10.3389","volume":"9","author":[{"given":"Jordi","family":"Ribas-Maynou","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ariadna","family":"Delgado-Berm\u00fadez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Estela","family":"Garcia-Bonavila","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Elisabeth","family":"Pinart","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marc","family":"Yeste","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sergi","family":"Bonet","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2021,6,14]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.3390\/IJMS21113882","article-title":"The impact of single- and double-strand DNA breaks in human spermatozoa on assisted reproduction.","volume":"21","author":"Agarwal","year":"2020","journal-title":"Int. J. Mol. Sci."},{"key":"B2","doi-asserted-by":"publisher","first-page":"401","DOI":"10.1016\/s1472-6483(10)60602-5","article-title":"Increased concentrations of the oxidative DNA adduct 7,8-dihydro-8-oxo-2-deoxyguanosine in the germ-line of men with type 1 diabetes.","volume":"16","author":"Agbaje","year":"2008","journal-title":"Reprod. Biomed. Online"},{"key":"B3","doi-asserted-by":"publisher","first-page":"3","DOI":"10.1093\/molehr\/gap059","article-title":"On the possible origins of DNA damage in human spermatozoa.","volume":"16","author":"Aitken","year":"2010","journal-title":"Mol. Hum. Reprod."},{"key":"B4","doi-asserted-by":"publisher","first-page":"938","DOI":"10.1093\/humrep\/dew030","article-title":"HT-COMET: a novel automated approach for high throughput assessment of human sperm chromatin quality.","volume":"31","author":"Albert","year":"2016","journal-title":"Hum. Reprod."},{"key":"B5","doi-asserted-by":"publisher","first-page":"824","DOI":"10.1093\/humrep\/dep465","article-title":"Sperm DNA: organization, protection and vulnerability: from basic science to clinical applications\u2013a position report.","volume":"25","author":"Barratt","year":"2010","journal-title":"Hum. Reprod."},{"key":"B6","doi-asserted-by":"publisher","first-page":"2075","DOI":"10.1016\/s0140-6736(00)05179-5","article-title":"Conventional in-vitro fertilisation versus intracytoplasmic sperm injection for the treatment of non-male-factor infertility: a randomised controlled trial.","volume":"357","author":"Bhattacharya","year":"2001","journal-title":"Lancet"},{"key":"B7","doi-asserted-by":"publisher","first-page":"728","DOI":"10.1016\/j.theriogenology.2007.12.004","article-title":"Sperm chromatin structure integrity in liquid stored boar semen and its relationships with field fertility.","volume":"69","author":"Boe-Hansen","year":"2008","journal-title":"Theriogenology"},{"key":"B8","doi-asserted-by":"publisher","first-page":"93","DOI":"10.1016\/j.theriogenology.2019.05.043","article-title":"An update on boar semen assessments by flow cytometry and CASA.","volume":"137","author":"Boe-Hansen","year":"2019","journal-title":"Theriogenology"},{"key":"B9","doi-asserted-by":"publisher","first-page":"483","DOI":"10.1016\/j.fertnstert.2019.04.029","article-title":"Sperm DNA fragmentation is correlated with poor embryo development, lower implantation rate, and higher miscarriage rate in reproductive cycles of non\u2013male factor infertility.","volume":"112","author":"Borges","year":"2019","journal-title":"Fertil. Steril."},{"key":"B10","doi-asserted-by":"publisher","DOI":"10.1111\/and.13012","article-title":"Reactive oxygen species impact on sperm DNA and its role in male infertility.","volume":"50","author":"Bui","year":"2018","journal-title":"Andrologia"},{"key":"B11","doi-asserted-by":"publisher","first-page":"1401","DOI":"10.1093\/humrep\/deh280","article-title":"The predictive value of sperm chromatin structure assay (SCSA) parameters for the outcome of intrauterine insemination, IVF and ICSI.","volume":"19","author":"Bungum","year":"2004","journal-title":"Hum. Reprod."},{"key":"B12","doi-asserted-by":"publisher","first-page":"699","DOI":"10.1016\/j.fertnstert.2018.11.035","article-title":"Double-stranded sperm DNA damage is a cause of delay in embryo development and can impair implantation rates.","volume":"111","author":"Casanovas","year":"2019","journal-title":"Fertil. Steril."},{"key":"B13","doi-asserted-by":"publisher","first-page":"S621","DOI":"10.21037\/tau.2017.06.25","article-title":"The correct interpretation of sperm DNA fragmentation test.","volume":"6","author":"Cho","year":"2017","journal-title":"Transl. Androl. Urol."},{"key":"B14","doi-asserted-by":"publisher","first-page":"421","DOI":"10.1111\/andr.12689","article-title":"Investigating the level of DNA double-strand break in human spermatozoa and its relation to semen characteristics and IVF outcome using phospho-histone H2AX antibody as a biomarker.","volume":"8","author":"Coban","year":"2020","journal-title":"Andrology"},{"key":"B15","doi-asserted-by":"publisher","first-page":"354","DOI":"10.1111\/j.1365-2605.2007.00781.x","article-title":"Alkali-labile sites in sperm cells from Sus and Ovis species.","volume":"31","author":"Cort\u00e9s-Guti\u00e9rrez","year":"2008","journal-title":"Int. J. Androl."},{"key":"B16","doi-asserted-by":"publisher","DOI":"10.3389\/fgene.2014.00404","article-title":"Interpreting sperm DNA damage in a diverse range of mammalian sperm by means of the two-tailed comet assay.","volume":"5","author":"Cort\u00e9s-Guti\u00e9rrez","year":"2014","journal-title":"Front. Genet."},{"key":"B17","doi-asserted-by":"publisher","DOI":"10.1111\/and.13263","article-title":"Sperm DNA fragmentation index influences assisted reproductive technology outcome: a systematic review and meta-analysis combined with a retrospective cohort study.","volume":"51","author":"Deng","year":"2019","journal-title":"Andrologia"},{"key":"B18","doi-asserted-by":"publisher","first-page":"633","DOI":"10.1071\/RD10269","article-title":"Differential resistance of mammalian sperm chromatin to oxidative stress as assessed by a two-tailed comet assay.","volume":"23","author":"Enciso","year":"2011","journal-title":"Reprod. Fertil. Dev."},{"key":"B19","doi-asserted-by":"publisher","first-page":"308","DOI":"10.1016\/j.theriogenology.2005.05.044","article-title":"A new method to analyze boar sperm DNA fragmentation under bright-field or fluorescence microscopy.","volume":"65","author":"Enciso","year":"2006","journal-title":"Theriogenology"},{"key":"B20","doi-asserted-by":"crossref","first-page":"609","DOI":"10.1016\/S1472-6483(10)60003-X","article-title":"A two-tailed Comet assay for assessing DNA damage in spermatozoa.","volume":"18","author":"Enciso","year":"2009","journal-title":"Reprod. Biomed. Online"},{"key":"B21","doi-asserted-by":"publisher","first-page":"209","DOI":"10.1016\/j.mrrev.2003.09.007","article-title":"The use of the alkaline comet assay with lymphocytes in human biomonitoring studies.","volume":"566","author":"Faust","year":"2004","journal-title":"Mutat. Res."},{"key":"B22","doi-asserted-by":"publisher","first-page":"833","DOI":"10.1016\/j.fertnstert.2004.11.089","article-title":"Simple determination of human sperm DNA fragmentation with an improved sperm chromatin dispersion test.","volume":"84","author":"Fern\u00e1ndez","year":"2005","journal-title":"Fertil. Steril."},{"key":"B23","doi-asserted-by":"publisher","first-page":"77","DOI":"10.1016\/S0027-5107(00)00079-8","article-title":"DNA breakage detection-FISH (DBD-FISH) in human spermatozoa: technical variants evidence different structural features.","volume":"453","author":"Fern\u00e1ndez","year":"2000","journal-title":"Mutat. Res. Fundam. Mol. Mech. Mutagen."},{"key":"B24","doi-asserted-by":"publisher","DOI":"10.1111\/j.1439-0531.2009.01537.x","article-title":"Comparison of post-thaw DNA integrity of boar spermatozoa assessed with the neutral comet assay and Sperm-Sus Halomax test kit.","volume":"45","author":"Fraser","year":"2010","journal-title":"Reprod. Domest. Anim."},{"key":"B25","doi-asserted-by":"publisher","first-page":"317","DOI":"10.1016\/j.anireprosci.2006.06.003","article-title":"Effect of different procedures of ejaculate collection, extenders and packages on DNA integrity of boar spermatozoa following freezing-thawing.","volume":"99","author":"Fraser","year":"","journal-title":"Anim. Reprod. Sci."},{"key":"B26","doi-asserted-by":"publisher","first-page":"248","DOI":"10.1016\/j.theriogenology.2007.05.001","article-title":"Is there a relationship between the chromatin status and DNA fragmentation of boar spermatozoa following freezing-thawing?","volume":"68","author":"Fraser","year":"","journal-title":"Theriogenology"},{"key":"B27","doi-asserted-by":"publisher","first-page":"119","DOI":"10.1016\/j.anireprosci.2014.04.010","article-title":"Post-thaw sperm characteristics following long-term storage of boar semen in liquid nitrogen.","volume":"147","author":"Fraser","year":"2014","journal-title":"Anim. Reprod. Sci."},{"key":"B28","doi-asserted-by":"publisher","first-page":"130","DOI":"10.1016\/j.mrrev.2019.04.002","article-title":"The comet assay in animal models: from bugs to whales - (Part 2 Vertebrates).","volume":"781","author":"Gajski","year":"2019","journal-title":"Mutat. Res."},{"key":"B29","doi-asserted-by":"publisher","first-page":"276","DOI":"10.1095\/biolreprod53.2.276","article-title":"Viability assessment of mammalian sperm using SYBR-14 and propidium iodide.","volume":"53","author":"Garner","year":"1995","journal-title":"Biol. Reprod."},{"key":"B30","doi-asserted-by":"publisher","first-page":"445","DOI":"10.1016\/j.theriogenology.2004.09.024","article-title":"Flow cytometric evaluation of sperm parameters in relation to fertility potential.","volume":"63","author":"Gillan","year":"2005","journal-title":"Theriogenology"},{"key":"B31","doi-asserted-by":"publisher","first-page":"812","DOI":"10.1038\/aja.2013.74","article-title":"Can DNA fragmentation of neat or swim-up spermatozoa be used to predict pregnancy following ICSI of fertile oocyte donors?","volume":"15","author":"Gos\u00e1lvez","year":"2013","journal-title":"Asian J. Androl."},{"key":"B32","doi-asserted-by":"publisher","first-page":"583","DOI":"10.1111\/j.1365-2605.2006.00699.x","article-title":"Differences in SCSA outcome among boars with different sperm freezability.","volume":"29","author":"Hern\u00e1ndez","year":"2006","journal-title":"Int. J. Androl."},{"key":"B33","doi-asserted-by":"publisher","first-page":"257","DOI":"10.1016\/j.cryobiol.2008.09.004","article-title":"Effects of different extenders on DNA integrity of boar spermatozoa following freezing-thawing.","volume":"57","author":"Hu","year":"2008","journal-title":"Cryobiology"},{"key":"B34","doi-asserted-by":"publisher","DOI":"10.3389\/fgene.2015.00266","article-title":"The comet assay: past, present, and future.","volume":"6","author":"Langie","year":"2015","journal-title":"Front. Genet."},{"key":"B35","doi-asserted-by":"publisher","first-page":"163","DOI":"10.1093\/mutage\/gem052","article-title":"Using the alkaline comet assay in prognostic tests for male infertility and assisted reproductive technology outcomes.","volume":"23","author":"Lewis","year":"2008","journal-title":"Mutagenesis"},{"key":"B36","doi-asserted-by":"publisher","first-page":"33","DOI":"10.1007\/s00441-005-1097-5","article-title":"DNA damage to spermatozoa has impacts on fertilization and pregnancy.","volume":"322","author":"Lewis","year":"2005","journal-title":"Cell Tissue Res."},{"key":"B37","doi-asserted-by":"publisher","first-page":"201","DOI":"10.3109\/14647273.2010.528823","article-title":"Clinical implications of sperm DNA damage.","volume":"13","author":"Lewis","year":"2010","journal-title":"Hum. Fertil."},{"key":"B38","doi-asserted-by":"publisher","first-page":"2","DOI":"10.1111\/j.1365-2605.2009.01042.x","article-title":"The TUNEL assay consistently underestimates DNA damage in human spermatozoa and is influenced by DNA compaction and cell vitality: development of an improved methodology.","volume":"34","author":"Mitchell","year":"2011","journal-title":"Int. J. Androl."},{"key":"B39","doi-asserted-by":"publisher","DOI":"10.3390\/jcm9051341","article-title":"DNA fragmentation in viable and non-viable spermatozoa discriminates fertile and subfertile subjects with similar accuracy.","volume":"9","author":"Muratori","year":"2020","journal-title":"J. Clin. Med."},{"key":"B40","doi-asserted-by":"publisher","first-page":"203","DOI":"10.1093\/molehr\/gah029","article-title":"Structure of human sperm DNA and background damage, analysed by in situ enzymatic treatment and digital image analysis.","volume":"10","author":"Muriel","year":"2004","journal-title":"Mol. Hum. Reprod."},{"key":"B41","doi-asserted-by":"publisher","first-page":"160","DOI":"10.1111\/rda.13322","article-title":"Sperm DNA integrity in Landrace and Duroc boar semen and its relationship to litter size.","volume":"54","author":"Myromslien","year":"2019","journal-title":"Reprod. Domest. Anim."},{"key":"B42","doi-asserted-by":"publisher","first-page":"F93","DOI":"10.1530\/REP-17-0374","article-title":"Intracytoplasmic sperm injection: state of the art in humans.","volume":"154","author":"Palermo","year":"2017","journal-title":"Reproduction"},{"key":"B43","doi-asserted-by":"publisher","first-page":"235","DOI":"10.1016\/j.anireprosci.2010.01.002","article-title":"Dynamics of sperm DNA fragmentation in the swine: ejaculate and temperature effects.","volume":"119","author":"P\u00e9rez-Llano","year":"2010","journal-title":"Anim. Reprod. Sci."},{"key":"B44","doi-asserted-by":"publisher","DOI":"10.3390\/genes10020105","article-title":"Single and double strand sperm DNA damage: different reproductive effects on male fertility.","volume":"10","author":"Ribas-Maynou","year":"2019","journal-title":"Genes"},{"key":"B45","doi-asserted-by":"publisher","DOI":"10.3389\/fcell.2021.669182","article-title":"Species-specific differences in sperm chromatin decondensation between eutherian mammals underlie distinct lysis requirements.","volume":"9","author":"Ribas-Maynou","year":"","journal-title":"Front. Cell Dev. Biol."},{"key":"B46","doi-asserted-by":"publisher","first-page":"652","DOI":"10.1093\/humrep\/der461","article-title":"Alkaline and neutral Comet assay profiles of sperm DNA damage in clinical groups.","volume":"27","author":"Ribas-Maynou","year":"","journal-title":"Hum. Reprod."},{"key":"B47","doi-asserted-by":"publisher","first-page":"715","DOI":"10.1111\/j.2047-2927.2013.00111.x","article-title":"Comprehensive analysis of sperm DNA fragmentation by five different assays: TUNEL assay, SCSA, SCD test and alkaline and neutral Comet assay.","volume":"1","author":"Ribas-Maynou","year":"2013","journal-title":"Andrology"},{"key":"B48","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0044679","article-title":"Double stranded sperm DNA breaks, measured by Comet assay, are associated with unexplained recurrent miscarriage in couples without a female factor.","volume":"7","author":"Ribas-Maynou","year":"","journal-title":"PLoS One"},{"key":"B49","doi-asserted-by":"publisher","first-page":"83","DOI":"10.1111\/j.2047-2927.2013.00158.x","article-title":"Human semen cryopreservation: a sperm DNA fragmentation study with alkaline and neutral Comet assay.","volume":"2","author":"Ribas-Maynou","year":"2014","journal-title":"Andrology"},{"key":"B50","doi-asserted-by":"publisher","first-page":"57","DOI":"10.1111\/brv.12700","article-title":"Clinical implications of sperm DNA damage in IVF and ICSI: updated systematic review and meta-analysis.","volume":"49","author":"Ribas-Maynou","year":"","journal-title":"Biol. Rev."},{"key":"B51","doi-asserted-by":"publisher","DOI":"10.3390\/biology9070178","article-title":"The relationship between sperm oxidative stress alterations and IVF\/ICSI outcomes: a systematic review from nonhuman mammals.","volume":"9","author":"Ribas-Maynou","year":"2020","journal-title":"Biology"},{"key":"B52","doi-asserted-by":"publisher","first-page":"2908","DOI":"10.1093\/humrep\/des261","article-title":"The effect of sperm DNA fragmentation on miscarriage rates: a systematic review and meta-analysis.","volume":"27","author":"Robinson","year":"2012","journal-title":"Hum. Reprod."},{"key":"B53","doi-asserted-by":"publisher","first-page":"197","DOI":"10.1016\/0027-5107(93)90070-v","article-title":"DNA strand breaks and chromosomal aberrations induced by H2O2 and 60Co gamma-radiation.","volume":"289","author":"Rueff","year":"1993","journal-title":"Mutat. Res."},{"key":"B54","doi-asserted-by":"publisher","first-page":"1027","DOI":"10.1016\/j.fertnstert.2009.10.046","article-title":"Sperm DNA fragmentation: mechanisms of origin, impact on reproductive outcome, and analysis.","volume":"93","author":"Sakkas","year":"2010","journal-title":"Fertil. Steril."},{"key":"B55","doi-asserted-by":"publisher","DOI":"10.1186\/s12958-015-0035-y","article-title":"The relationship between sperm viability and DNA fragmentation rates.","volume":"13","author":"Samplaski","year":"2015","journal-title":"Reprod. Biol. Endocrinol."},{"key":"B56","doi-asserted-by":"publisher","DOI":"10.3389\/fgene.2015.00216","article-title":"The use of comet assay in plant toxicology: recent advances.","volume":"6","author":"Santos","year":"2015","journal-title":"Front. Genet."},{"key":"B57","doi-asserted-by":"publisher","first-page":"88","DOI":"10.1016\/j.theriogenology.2019.05.042","article-title":"New trends in production management in European pig AI centers.","volume":"137","author":"Schulze","year":"2019","journal-title":"Theriogenology"},{"key":"B58","doi-asserted-by":"publisher","first-page":"1380","DOI":"10.1016\/j.urology.2010.04.036","article-title":"TUNEL as a test for sperm DNA damage in the evaluation of male infertility.","volume":"76","author":"Sharma","year":"2010","journal-title":"Urology"},{"key":"B59","doi-asserted-by":"publisher","DOI":"10.1111\/and.12608","article-title":"Sperm DNA damage output parameters measured by the alkaline Comet assay and their importance.","volume":"49","author":"Simon","year":"","journal-title":"Andrologia"},{"key":"B60","doi-asserted-by":"publisher","first-page":"1594","DOI":"10.1093\/humrep\/deq103","article-title":"Clinical significance of sperm DNA damage in assisted reproduction outcome.","volume":"25","author":"Simon","year":"2010","journal-title":"Hum. Reprod."},{"key":"B61","doi-asserted-by":"publisher","first-page":"137","DOI":"10.1007\/978-1-62703-038-0_13","article-title":"Sperm DNA damage measured by comet assay.","volume":"927","author":"Simon","year":"2013","journal-title":"Methods Mol. Biol."},{"key":"B62","doi-asserted-by":"publisher","first-page":"652","DOI":"10.1016\/j.fertnstert.2010.08.019","article-title":"Sperm DNA damage measured by the alkaline Comet assay as an independent predictor of male infertility and in vitro fertilization success.","volume":"95","author":"Simon","year":"2011","journal-title":"Fertil. Steril."},{"key":"B63","doi-asserted-by":"publisher","first-page":"68","DOI":"10.1016\/j.rbmo.2012.09.019","article-title":"Sperm DNA damage has a negative association with live-birth rates after IVF.","volume":"26","author":"Simon","year":"2013","journal-title":"Reprod. Biomed. Online"},{"key":"B64","doi-asserted-by":"publisher","first-page":"80","DOI":"10.4103\/1008-682X.182822","article-title":"A systematic review and meta-analysis to determine the effect of sperm DNA damage on in vitro fertilization and intracytoplasmic sperm injection outcome.","volume":"19","author":"Simon","year":"","journal-title":"Asian J. Androl."},{"key":"B65","doi-asserted-by":"crossref","first-page":"184","DOI":"10.1016\/0014-4827(88)90265-0","article-title":"A simple technique for quantitation of low levels of DNA damage in individual cells.","volume":"175","author":"Singh","year":"1988","journal-title":"Exp. Cell Res."},{"key":"B66","doi-asserted-by":"publisher","first-page":"87","DOI":"10.1007\/s12522-013-0168-7","article-title":"Sperm DNA fragmentation assay by sperm chromatin dispersion (SCD): correlation between DNA fragmentation and outcome of intracytoplasmic sperm injection.","volume":"13","author":"Sivanarayana","year":"2014","journal-title":"Reprod. Med. Biol."},{"key":"B67","doi-asserted-by":"publisher","first-page":"678","DOI":"10.1111\/j.2047-2927.2014.00234.x","article-title":"Sperm DNA integrity testing: big halo is a good predictor of embryo quality and pregnancy after conventional IVF.","volume":"2","author":"Tandara","year":"2014","journal-title":"Andrology"},{"key":"B68","doi-asserted-by":"publisher","first-page":"843","DOI":"10.1016\/j.fertnstert.2011.07.356","article-title":"Oxidative deoxyribonucleic acid damage in sperm has a negative impact on clinical pregnancy rate in intrauterine insemination but not intracytoplasmic sperm injection cycles.","volume":"96","author":"Thomson","year":"2011","journal-title":"Fertil. Steril."},{"key":"B69","doi-asserted-by":"publisher","DOI":"10.1016\/j.anireprosci.2020.106350","article-title":"Computer Aided Sperm Analysis (CASA) in domestic animals: current status, three D tracking and flagellar analysis.","volume":"220","author":"van der Horst","year":"2020","journal-title":"Anim. Reprod. Sci."},{"key":"B70","doi-asserted-by":"publisher","first-page":"445","DOI":"10.1530\/REP-10-0176","article-title":"Sperm DNA fragmentation induced by DNAse I and hydrogen peroxide: an in vitro comparative study among different mammalian species.","volume":"140","author":"Villani","year":"2010","journal-title":"Reproduction"},{"key":"B71","doi-asserted-by":"publisher","first-page":"45","DOI":"10.1111\/j.1439-0531.2011.01836.x","article-title":"Assessment of storage effects in liquid preserved boar semen.","volume":"46","author":"Waberski","year":"2011","journal-title":"Reprod. Domest. Anim."},{"key":"B72","doi-asserted-by":"publisher","first-page":"2","DOI":"10.1016\/J.THERIOGENOLOGY.2019.05.030","article-title":"Application of preserved boar semen for artificial insemination: past, present and future challenges.","volume":"137","author":"Waberski","year":"2019","journal-title":"Theriogenology"},{"key":"B73","doi-asserted-by":"publisher","first-page":"30","DOI":"10.1093\/molehr\/gap080","article-title":"Function of sperm chromatin structural elements in fertilization and development.","volume":"16","author":"Ward","year":"2010","journal-title":"Mol. Hum. Reprod."},{"key":"B74","doi-asserted-by":"publisher","first-page":"929","DOI":"10.1016\/j.theriogenology.2013.01.008","article-title":"Good and bad freezability boar ejaculates differ in the integrity of nucleoprotein structure after freeze-thawing but not in ROS levels.","volume":"79","author":"Yeste","year":"","journal-title":"Theriogenology"},{"key":"B75","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0090887","article-title":"The increase in phosphorylation levels of serine residues of protein HSP70 during holding time at 17\u00b0C is concomitant with a higher cryotolerance of boar spermatozoa.","volume":"9","author":"Yeste","year":"2014","journal-title":"PLoS One"},{"key":"B76","doi-asserted-by":"publisher","first-page":"1036","DOI":"10.1071\/RD12230","article-title":"Reduced glutathione and procaine hydrochloride protect the nucleoprotein structure of boar spermatozoa during freeze-thawing by stabilising disulfide bonds.","volume":"25","author":"Yeste","year":"","journal-title":"Reprod. Fertil. Dev."},{"key":"B77","doi-asserted-by":"publisher","first-page":"802","DOI":"10.1002\/mrd.22840","article-title":"Artificial insemination with frozen-thawed boar sperm.","volume":"84","author":"Yeste","year":"2017","journal-title":"Mol. Reprod. Dev."},{"key":"B78","doi-asserted-by":"publisher","first-page":"1004","DOI":"10.1002\/mrd.22875","article-title":"Genetic control of complex traits, with a focus on reproduction in pigs.","volume":"84","author":"Zak","year":"2017","journal-title":"Mol. Reprod. Dev."}],"container-title":["Frontiers in Cell and Developmental Biology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fcell.2021.675973\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,6,14]],"date-time":"2021-06-14T02:58:17Z","timestamp":1623639497000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fcell.2021.675973\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,6,14]]},"references-count":78,"alternative-id":["10.3389\/fcell.2021.675973"],"URL":"https:\/\/doi.org\/10.3389\/fcell.2021.675973","relation":{},"ISSN":["2296-634X"],"issn-type":[{"value":"2296-634X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,6,14]]},"article-number":"675973"}}