2.2 Cloning the target sequence
The upstream region of the carotenoid cleavage dioxygenase 8 (CCD8) gene
[13] from Z. mays L. cv M37W was amplified by PCR from
genomic DNA prepared with the EchoLUTION Plant DNA Kit (BioEcho Life
Sciences GmbH, Cologne, Germany) using the primer pair Zm_CCD8_for
5’-CAGGCAGCAAACGCTCACCACCAGGC and Zm_CCD8_rev
5’-CGACGAAGCCATAGTGGGAGACATGGCG. The amplicon was cloned into pJET1.2
(CloneJET PCR Cloning Kit, Thermo Fisher Scientific) and verified by
Sanger sequencing. For subsequent assays, high-quality plasmid DNA was
prepared from E. coli harboring pJET_CCD8 using the NucleoBond
Xtra Midi Plus kit (Macherey & Nagel, Düren, Germany).