Objective <p>This study aims to evaluate the association between ultrasound soft markers and fetal chromosomal abnormalities and to compare the diagnostic efficacy of karyotype analysis versus chromosomal microarray analysis (CMA) for prenatal testing strategy optimization.</p> Materials and methods <p>A retrospective review was conducted on 622 cases receiving prenatal diagnosis for abnormal ultrasound soft markers at our center over three years. All cases underwent chromosomal karyotype analysis and CMA testing. The differences between the results of these two tests, as well as the correlation between genetic testing results and abnormal ultrasound soft markers, were analyzed. Additionally, the pregnancy outcomes and postnatal phenotypes of all cases were monitored.</p> Results <p>The overall prevalence of chromosomal abnormalities was 11.41% (71/622). Echogenic intracardiac focus (<i>P</i> = 0.012) and multiple soft markers (<i>P</i> &lt; 0.001) exhibited a higher correlation with chromosomal abnormalities, with the latter showing a particularly strong association with aneuploidy (<i>P</i> &lt; 0.001). Karyotype analysis identified 63 chromosomal abnormalities, while CMA detected 65, with discordant results observed in 18 cases. Among the cases with chromosomal abnormalities, 11 resulted in live births, and follow-up at ages 3–5 revealed no abnormal phenotypes.</p> Conclusion <p>Prenatal genetic diagnosis is strongly recommended for pregnant women presenting with ultrasound soft markers, particularly multiple markers. Concurrent CMA and karyotype analysis are advocated to minimize the risk of missing pathogenic variants.</p>