Preparation of Polyclonal Antibody of Heart Development Marker Gene Kruppel in Drosophila

Kruppel plays a very important role in development of heart.Obtianing the Kruppel protein and it's antibody is the first step to detect the function of Kruppel.After the sequence of Kruppel was analyzed and the primer was designed,the part of encoding sequence of drosophila Kruppel was amplified with PCR from cDNA of drosophila heart,and inserted into pET-28a vector in frame.The competent cells of host strain of rosetta were transformed by the recombinant plasmid(pET-28a-Kruppel).Expression of the target protein was induced with IPTG and purified by Ni-NTA affinity chromatography under denaturing conditions.Then the New Zealand white rabbits was immuned with purified recombinant protein to generate antibody.The antibody titer was identified by Western blot.