Root-knot nematodes (RKN) and root-lesion nematodes are plant endoparasites. The Meloidogyne spp. of RKN can infect more than 3000 plant species and are responsible for annual losses of around $100 billion worldwide. Non-chemical control methods are often slow and less effective, whereas the efficient chemical control methods can be toxic to food supplies and the environment. Similar to insecticides that lose efficacy over time, emerging resistance towards chemical nematicides by nematodes could be a rising concern in the near future. Cyclotides are small cyclic mini-peptides distributed among five plant families. They are ~30 amino acids in size with three disulfide bonds connected in a knotted topology to form the cystine knot motif and a cyclic peptide backbone, together referred to as cyclic cystine knot (CCK). Kalata B1 (kB1) is a plant-derived cyclotide that had reported nematicidal activity against the larval stage of gastrointestinal nematode parasites of sheep. Cyclotide kB1 was expressed in transgenic ex-vitro hairy roots of peanut explants using a plasmid encoding the multiple genes involved for its production. Another plasmid with a green fluorescent protein (GFP) was used as the control for these experiments. The detection of kB1 in roots could be done by using MALDI and the GFP positive roots could be checked simply with a blue torch. This production system could be further improved by assembling a recombinant plasmid combining the cyclotide kB1 production genes with the GFP gene. This will help in easy and quick selection of positive hairy root lines emerging from the explant. Subsequently, these roots will be exposed to RKN in the soil to study the effects and interaction of kB1 on RKN.