FASTA format. This sequence may be either pasted in or a sequence ﬁle may be uploaded. AMOD then performs TIS calculations on all entered sequences and asks users whether BLAST comparisons are desired. Following the com- putation or bypass of BLAST comparisons, users are prompted to select a single input sequence against which morpholino design will proceed. Once a user has deﬁned a subsequence, reviewed the putative morpholinos, and identiﬁed an accept- able oligonucleotide, the morpholino sequence characteristics Figure 3. Illustrates intra-strand binding approximation from a morpholino self-to-self alignment. In this figure, the total binding score is 30: 18 from six G– C bonds and 12 from six A–T bonds. Nucleic Acids Research, 2005, Vol. 33, Web Server issue W509 are written to an output ﬁle. The user may then analyze another sequence from the input sequence set or exit the program. AMOD application AMOD was used to design morpholinos for the University of Minnesota’s functional genomics screen in zebraﬁsh. This screen utilizes comparative genomic data mining strategies to identify conservation of higher-order biological processes among vertebrates. Use of morpholinos allows high-throughput rates of genetic analysis without the inherent redundancy of forward-genetic approaches that typically result in multiple mutations within the same gene. The screen has analyzed 255 gene sequences to date, using 189 morpholinos targeting 157 unique genes were designed. During AMOD’s use, the program’s functionality and user interface were evaluated and improved. The positive results of the ongoing functional genomics morpholino screen validate the use of morpholinos for this approach (27) and demonstrate the capacity of AMOD to Figure 4. Oligonucleotide selection phase. (A) All intra-strand binding alignments are reported, and the alignments yielding the maximum global intra-strand binding and the maximum intra-strand binding constrained to a four-base window are noted. (B) AMOD uses a second set of BLAST comparisons between select morpholinos and genomic sequences or the transcriptome to identify genes that could be targeted by the selected morpholino oligonucleotides.