Correlation between the recombinant gene dosage and the expression levels of the aquaporins. Diagrams showing the μg/mL zeocin selection level and the AOX1 TT copy numbers ± SD of clones carrying the HsAQP8 (A), HsAQP5 (B), SoPIP1;2 (C) and AtSIP1;1 (D) constructs. For the SoPIP1;2 construct, two clones were recovered from the 2000 μg/mL zeocin selection. This higher antibiotic resistance is indicated by a darker colour in the diagram. The AOX1 TT copy numbers include the native AOX1 TT sequence and the sequences derived from the incorporated plasmids. For the X-33 wild type strain, included as a standard, where no vector derived AOX1 TT sequences are present, a 1:1 ratio between the AOX1 TT sequence and the internal reference sequence is expected and a value of 1 ± SD is calculated. By subtracting this value from the AOX1 TT copy numbers obtained for the recombinant clones, the aquaporin gene copy numbers were subsequently calculated (see methods and Figure 6). Included are also western blots showing recombinant expression levels and diagrams summarising the densitometric quantification of each of the lanes with total protein (black bars) and degradation products (grey bars) indicated. Values are given as Arbitrary Intensity Units (AIU:s). There is a correlation between aquaporin gene copy number and antibiotic selection level and between aquaporin expression and aquaporin gene copy number.