Team:Bilkent UNAM Turkey/Experiment

We ordered our genes. We optimized codon usage because of gene taken from Enterobacter cloacae. Codon Usage . We optimized our NFSI gene according to C.reinhardtii codon bias table provided by DNA 2.0. This program enables to change synonymous codons without changing amino acid sequence. You can also check the restriction sites while optimizing codons. We also avoided from forbidden restriction sites (EcoRI, XbaI, SpeI, PstI) therefore; we have chosen second frequently used codons for 34., 35., 38. and 45. amino acids. Then we did restriction digestion and gel electrophoresis.
We repeated this step a lot because of failuire.I put right result







We used standard gel purification kit Our ligation style is really similar to iGEM's one.
Our one;

insert volume(µl):X

Cut Vector Volume(µl):X

DEPC-water(µl):8,5-2X

T4 DNA ligase(µl):0,5

Ligation Buffer(µl):1

Total Volume(µl):10

We transformed our plasmid into E.coli with a protocol.
We used invitrogen purification kit to get our plasmid. We sent genes to sequencing and sequencing data should be available on their link.