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Background and Purpose: Genomic nursing education requires a genomic literacy assessment supported by evidence of reliability and validity. This study applied psychometric analyses to provide support for the Genomic Nursing Concept Inventory (GNCI). Methods: Over nine semesters, baccalaureate nursing students (N = 1,065) completed the GNCI on the first and last days of genomics instruction. Psychometric analyses assessed scale and item performance pre- and post-instruction. Results: Exploratory factor analysis supported scale unidimensionality and identified items with low pattern coefficients. Analyses supported test–retest and internal consistency reliability and criterion validity. Scale difficulty decreased by 28% from pre- to post-instruction. Underperforming items were identified for further inventory refinement. Conclusions: Findings support use of the GNCI to measure learning needs pre-instruction and learning gains post-instruction. Data also inform planned inventory revision. [ABSTRACT FROM AUTHOR]