The interaction between CFTR mutants and/or polymorphisms results in different CFTR channel functions; which could explain why apparently normal CFTR genes cause disease. Both TM 12 and the intra-cytoplasmic stretch connecting TM 12 and NBD2 probably contribute to form the CFTR pore but may not contribute to anion selectivity. The majority of the phosphorylation sites identified so far occur within the R domain. Phosphorylation of PKA sites present in this domain is the initial event in channel activation and the degree of phosphorylation determines the affinity of both nucleotide binding domains for ATP. One major goal of current research is to understand how phosphorylation of multiple R domain serines by PKA effectively regulates CFTR channel function. Results obtained in this study, using constructs mutated at amino acids located in exon 13, and deletion constructs, indicated that the R domain indeed plays a crucial role in regulating the CFTR channel. The regulation efficiency is, at least in part, related to electric charge. Replacing the negatively charged glutamic acids to positively charged lysine at positions 822 and 826 in the C-terminal part of R domain resulted in an almost complete loss of CFTR channel activity. On the other hand, when a positively charged histidine was replaced by a more neutral amino acid at position 620 in the N-terminus of the R domain, CFTR channel activity was increased when compared with wildtype CFTR. Neither of these mutants affected open pore properties of the channel, since the single channel conductance and selectivity were similar for all mutants and the wildtype CFTR. Deletion of a small fragment in the C-terminus of the R domain (amino acids 780-830) resulted in a complete loss of CFTR channel activity, which could be restored by replacement of that fragment by the mini MDR1 linker domain, which has a different amino acid sequence but similar number of amino acids and phosphorylation sites. This result implie