In Pakistan, cotton crop contributes 23% to the GDP and its massive export provides 60% of the total profit in trading business. Unfortunately, the Gemini viruses destroy the cotton crop at an alarming rate. Now, the CLCuVs causes the Cotton Leaf Curl Virus Disease in cotton crop and reduce its productivity. This viral attack on cotton plant resulted 5 billion US dollars loss to the Pakistan. Well, in that regard, some conventional methodologies were used like the plant breeding and specific RNA editing. At the same time, biotechnology introduced some very attractive techniques which carry the massive potential for the eradication of this disease. These techniques include the ZFNs, TALENs and CRISPR/Cas9. In my research, CRISPR methodology was adopted because of its marvelous efficiency of site specific mutagenesis. The conserved locations of the rep gene of the different CLCuVs were identified as the target sites. These specific sites provided the information for the establishment of the 3 gRNAs. The single expression vector pHSE-401 having the multiple guided RNAs and 1Cas9 were cloned. The specific cotton variety coker-312 was used. The transformation of the vector was performed by hypocotyls excision of the cotton plant and delivery infection was done by agrobacterium EHA-105 Strain. Then, 1000 infectious hypocotyls were shifted on the MSB having respective antibiotics and then, on the regeneration media which converted the hypocotyls edges into the callus. Only two transgenic calli were obtained, the percentage of transgenic calli was 0.02% which were screened by PCR and run on gel. The bands of 200 bp confirmed the presence of the CRISPR/Cas9 construct in cotton calli.