incubated for 2h at 370C. The products were run in 1% agarose at 100V, using all the

incubation mixture.

 DNA sequencing and data processing. The concentration of the purified plasmid

DNA was determined prior to submission for sequencing. The DNA sequencing core

requires 1.5 Clg DNA for adequate processing. Cloned DNA sequences obtained were

then compared with nucleotide sequences in GenBank using the BLASTn tool provided

online (http://www.ncbi .nlm.nih.gov/BLAST). Multiple sequence comparisons were

done with SeqWeb, while two-sequence comparisons were done with the BLASTn 2.2.12

program.

 Results and discussion (part 1)

 Primer pairs 4 and 6 successfully amplified cDNA from catfish liver; while primer

pair 4 amplified cDNA from proximal intestine. The size of the amplicons were

approximately 300bp (pair 4) and 600bp (pair 6) in size, with the gel-clean up system

effectively removing primer dimers and other contamination (Figure 5-2). The controls

indicated that ligation and transformation of the plasmid into E.coli were successful.

Purified plasmid DNA was obtained from several colonies (Figure 5-3), which were

denoted as L1-L8 for the liver and 11-14 for the proximal intestine. The two bands seen in

these gels, did not represent contamination, as verified by the restriction digest of the

plasmid, which resulted in a band corresponding to the vector and one corresponding to

the smaller insert (Figure 5-4).