{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,18]],"date-time":"2026-03-18T06:59:45Z","timestamp":1773817185645,"version":"3.50.1"},"reference-count":36,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2026,1,7]],"date-time":"2026-01-07T00:00:00Z","timestamp":1767744000000},"content-version":"vor","delay-in-days":6,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2026,1,1]],"date-time":"2026-01-01T00:00:00Z","timestamp":1767225600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32172726"],"award-info":[{"award-number":["32172726"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32272872"],"award-info":[{"award-number":["32272872"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Reprod Domestic Animals"],"published-print":{"date-parts":[[2026,1]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>Livestock sex control is one of the core bioengineering technologies for improving quality and efficiency in modern animal husbandry, holding profound practical significance for sex\u2010limited livestock production systems. Precise identification of fetal sex at the early gestational stage constitutes a key prerequisite for achieving targeted sex regulation. The discovery of cell\u2010free fetal DNA (cffDNA) in the peripheral blood of pregnant cows has paved an important technical avenue for establishing a non\u2010invasive and high\u2010precision diagnostic system for early fetal sex identification. In this study, plasma and serum samples collected from pregnant cows were used as experimental materials, and three protocols\u2014phenol\u2010chloroform extraction, heat\u2010based extraction, and a commercial kit specifically designed for isolating cffDNA from plasma\/serum\u2014were employed for cffDNA purification and isolation. Y\u2010chromosome\u2010specific genes (either the TSPY or SRY gene) were selected as molecular markers, and optimised detection systems were established by integrating polymerase chain reaction (PCR), real\u2010time quantitative PCR (RT\u2010qPCR), and loop\u2010mediated isothermal amplification (LAMP) techniques. A systematic comparison was conducted to evaluate the efficacy and accuracy of different cffDNA extraction methods combined with various amplification technologies for fetal sex identification in both early and late stages of gestation. The actual calving outcomes were used as the standard for validation. The results demonstrated that the quality of cffDNA templates extracted by the commercial kit method was significantly superior to that obtained by the heat\u2010based and phenol\u2010chloroform methods, with the corresponding sex identification accuracy reaching the highest level. Notably, the LAMP technique exhibited unique advantages in detecting fetal sex in extremely early gestational samples (at 1\u20132\u2009months of pregnancy). Characterised by its simplicity of operation, rapid reaction kinetics, and elimination of the need for sophisticated instrumentation, LAMP is particularly well\u2010suited for on\u2010site large\u2010scale rapid primary screening of fetal sex in livestock farms. It enables the efficient exclusion of male foetuses within a short timeframe, thereby substantially improving the efficiency of breeding selection. Based on the aforementioned findings, this study proposes a combined detection model of \u201cLAMP\u2010based primary screening plus PCR\u2010based confirmation\u201d, which can effectively balance detection efficiency and identification accuracy. The research outcomes provide empirical data and methodological references for constructing a non\u2010invasive, early\u2010stage, and high\u2010precision technical system for fetal sex identification in dairy cows. This holds great value for promoting the implementation of precise early reproductive management in dairy farms and enhancing the economic benefits of the livestock industry.<\/jats:p>","DOI":"10.1111\/rda.70164","type":"journal-article","created":{"date-parts":[[2026,1,7]],"date-time":"2026-01-07T12:48:27Z","timestamp":1767790107000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Study of Methods for Early Fetal Sex Identification Using Cell\u2010Free Fetal\n                    <scp>DNA<\/scp>\n                    in the Peripheral Blood of Pregnant Cows"],"prefix":"10.1111","volume":"61","author":[{"given":"Yuxin","family":"Luo","sequence":"first","affiliation":[{"name":"College of Animal Science Jilin University  Changchun China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yaling","family":"Mi","sequence":"additional","affiliation":[{"name":"Hunan Institute of Animal and Veterinary Science  Changsha China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jinxin","family":"Zong","sequence":"additional","affiliation":[{"name":"College of Animal Science Jilin University  Changchun China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yang","family":"Luo","sequence":"additional","affiliation":[{"name":"Hunan Institute of Animal and Veterinary Science  Changsha China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lu","family":"Chen","sequence":"additional","affiliation":[{"name":"College of Animal Science Jilin University  Changchun China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kangle","family":"Yi","sequence":"additional","affiliation":[{"name":"Hunan Institute of Animal and Veterinary Science  Changsha China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Chunjin","family":"Li","sequence":"additional","affiliation":[{"name":"College of Animal Science Jilin University  Changchun China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xu","family":"Zhou","sequence":"additional","affiliation":[{"name":"College of Animal Science Jilin University  Changchun 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