Introduction: Polycystic ovary syndrome (POCS) is a mystery disorder with mysterious multiple players characterized by their mystic effects on disease pathophysiology resulting in various phenotypic pictures among the PCOS population. The Luteinizing hormone beta subunit (LH-B) (protein ID P01229) is a gonadotropin hormone secreted from the anterior pituitary belongs to the glycoprotein family, mapped on (chr19p13.3) and consists of three exons. It has a central role in promoting ovulation via stimulation of ovarian steroidogenesis. Objectives: This is a prospective laboratory-based cross-sectional study to determine genetic mutations associate with polycystic ovary syndrome (PCOS) among (30) Sudanese families ((cases n=35 families, 90 females, and (controls n= 11 families, 30 females) in Khartoum State, Sudan. Methods: Quantitative Enzyme-Linked Immuno-Sorbent Assay (ELISA), enzymatic methods, and polymerase chain reaction (PCR) used to analyze both the biochemical parameters and polymorphism detection followed by Sanger sequencing for genotyping in addition to bioinformatics software for protein structure and function. Results: All the biochemical parameters levels of (FBG, LH, Testosterone, Insulin, and lipid profile) elevated from the control group were statistically significant except for the serum FSH (cases=5.4±4.6, controls=5.3±2.8) and PRL ((cases=12.4±8.2, controls=8.0±6.1)) which were statistically insignificant p=0.94 and p=0.06. After Sanger sequencing; (5) single nucleotide polymorphisms ((rs5030775, A18T), rs746167425, R22K), (rs1800447, W28R), (rs35270001, H30R) and (rs34349826, I35T)) located on (exon 2) of the LH beta gene was statistically significant with serum LH, Testosterone, and insulin levels among PCOS families. Conclusion: This is the first molecular identification of a family-based study in Sudan exploring the genetic of the LHB gene and interrelated its serum level with PCOS manifestation. The revelation of these mutations will give a clue to the genetic inheritance mode links and might explain the abnormal poor response of controlled ovarian stimulation tests in some PCOS women.