Construction of RNAi Expression Vector of NtSKP1 and Transformation in Tobacco

According to the sequence of SKP1 from Nicotiana tabacum var.Samsun NN(NtSKP1),two pairs of specific primers containing special restriction enzyme sites were designed.The target reverse and forward fragments(473 bp),were amplified and inserted into right and left side of intron in intermediated vector pHANNIBAL respectively.Then the target fragment about 3916 bp was cut by NotⅠ from recombinant intermediated vector,and cloned into binary plasmid pART27.Plant expression plasmid pART27-skp1sa,with inverted repeat DNA fragment of NtSKP1,was successfully constructed,and pART27-skp1sa was introduced into Agrobacterium tumefaciens LBA4404.Then transformed tobacco leaves,and the transgenic plants of tobacco were obtained.