A. B. C.

 0 123 4567 0 0 12 3 4 567 0 12 3 4 5678













Figure 5-25. PCR amplification of UGT using degenerate primers.

 In all cases, the same seven sets of primers were used under similar PCR
 conditions. cDNA templates were as follows: A, channel catfish AT45
 intestine; B, channel catfish AT17 intestine; C, largemouth bass liver (lane 8
 is similar to lane 6, but the PCR was run at a higher annealing temperature).
 Lane 0 represents 100kb ladder for (A) and 1kb ladder for (B) and (C).


 Conclusions and recommendations

 One full length UGT from catfish liver, together with an identical UGT from

catfish intestine, was successfully cloned. A partial sequence of another UGT from

catfish intestine was also cloned. By homology with mammalian UGTs, the full-length

catfish UGT clone appeared to be analogous to UGTIAl or UGTIA6. Expressing this

gene into suitable cells (e.g. V79 or HEK cells) and characterizing the resulting protein

should provide us with further information on glucuronidation in the catfish. Performing

enzyme assays with UGT-selective probes, such as bilirubin (UGTIAl) and serotonin

(UGTIA6) (Patten et al., 2001; Krishnaswami et al., 2003), would assist in such studies.

As Table 5-11 shows, there are several potential antigenic sites on the predicted protein

sequence that may be exploited to design specific anti-UGT antibodies which would