FIG. 6.

RT-PCR reveals polycistronic mRNAs. Reverse transcriptase (RT) PCR was performed using 1 μg of FZB42 RNA isolated from stationary phase using a commercially available kit. PCR products were analyzed in comparison to a negative control lacking reverse transcriptase (−RT). (A) All genes in the putative PZN cluster are transcribed under the culturing conditions employed. Cells (1 ml at an OD₆₀₀ of 1.0) were removed from a stationary culture growing at 37°C 24 h after inoculation. Total RNA was isolated and converted to cDNA by RT-PCR. cDNA (500 ng), excluding pznE and pznL (750 ng each), was added to each reaction mixture. Gene fragments were then amplified using specific primers and PCR. Amplicons were assessed by separation on 1.1% agarose gels containing ethidium bromide and visualized by UV illumination. (B) Amplification of adjacent pzn genes reveals polycistronic mRNA. Junction I-J did not reveal a significant band; junction E-L was visible under extreme contrast (data not shown). All amplicons migrate on the basis of their expected sizes. Numbers at left represent DNA size standards.