Jew's mallow is used as a food source, medicinal plant and natural antioxidant agent as well as biofuel source. Its numerous beneficial properties made it a valuable crop worldwide. This study was carried out to investigate the morphology, biochemical genetic analysis and DNA barcoding for three different Jew's mallow (Corchorus olitorius) Egyptian landraces. They were collected from different location within Egypt namely, Fallahy (North), Seady (South) and Swain (West). Results revealed that, Siwan landrace had higher values for plant height, leaves number/plant, branches number/plant and fresh weight followed by Seady, then Fallahy. Biochemical values generally showed no significant differences between most of the measured elements. Mineral composition data showed numerical increase in the Siwan landrace in some minerals than others including Boron, Sodium, Magnesium, Aluminum, Calcium, Titanium, Chromium and Zinc, while Seady landrace recorded the highest values for Cobalt, Nickel, Copper, Indium and Bismuth. PCR was performed for the landraces along with two universal primers (RbcL-MatK). RbcL gene was amplified at 800-900 bp, while MatK gene was at 900 bp. Data was uploaded on DNAman program and partial sequences of RbcL and MatK genes were submitted to DDBJ Gene Bank with accession numbers of LC732565, LC732566 and LC732567 for RbcL gene and LC732049, LC732050 and LC732051 for MatK gene. This study showed that, the Siwan landraces was more unique from the other two landraces; furthermore, DNA barcoding using RbcL and MatK genes is an effective and sufficient tool in identifying different C. olitorius landraces in Egypt.