RT-PCR analysis of reassembled carotenoid cluster genes
E . coli harbouring recombinant pYS71, pYS69, or pYS76 was
grown in LB medium to exponential growth phase (12 h after inoculation).
The following primers were used for RT reactions: RT3 (crtZ ); RT4
(crtI ); and RT5 (crtY ). The following PCR primers were
used: PCR6f and PCR6r; PCR7f and PCR7r; and PCR8f and PCR8r
(Supplementary Table S3). Southern hybridisation and DNA sequencing were
carried out to confirm the RT-PCR products. As positive controls, pYS71,
pYS69, and pYS76 were used. As a negative control, PCR reactions with
the same primer sets were performed using RNA samples that had not been
reverse transcribed.