Cloning the MTP1 Gene from Chlamydomonas reinhardtii (2021)

Zinc plays an important role in processes such as cell growth and division and defensive immune systems. In humans, zinc metabolism may be linked to diseases such as diabetes. For this project we are placing the zinc transporter MTP1 from the green algae Chlamydomonas reinhardtii into the PET11 vector for expression in E. coli to produce the transporter so that this can be studied further. To accomplish this end, we use a polymerase chain reaction (PCR) to properly prepare the gene and plasmid PET11, followed by the NEBuilder HiFi DNA Assembly method to assemble them and place the new plasmid product into E. coli. We then identify bacteria potentially containing the desired plasmid product using colony PCR and plasmid isolation, endonuclease digests, and PAGE to analyze the plasmids for correct assembly.