Channel catfish (Ictalurus spp.) is an economically important species in freshwater aquaculture around the world and occupies a prominent position in the aquaculture industry of the United States. MicroRNAs (miRNAs) play important roles in the regulation of almost every biological process in eukaryotes; however, there is little information available concerning miRNAs in channel catfish. In this study, a small-RNA cDNA library was constructed from 10 tissues of channel catfish, and Solexa sequencing technology was used to perform high-throughput sequencing of the library. A total of 14,919,026 raw reads, representing 161,288 unique sequences, were obtained from the small-cDNA library. After comparing the small RNA sequences with the RFam database, 4,542,396 reads that represent 25,538 unique sequences were mapped to the genome sequence of zebrafish to perform distribution analysis and to screen for candidate miRNA genes. Subsequent bioinformatic analysis identified 237 conserved miRNAs and 45 novel miRNAs in the channel catfish. Stem-loop RT-PCR was applied to validate and profile the expression of the novel miRNAs in 10 tissues. Some novel miRNAs, such as ipu-miR-129b, ipu-miR-7562 and ipu-miR-7553, were expressed in all tissues examined. However, some novel miRNAs appear to be tissue specific. Ipu-miR-7575 is predominantly expressed in stomach. Ipu-miR-7147 and ipu-miR-203c are highly expressed in heart, but are relatively weakly expressed in other tissues. Based on sequence complementarity between miRNAs and mRNA targets, potential target sequences for the 45 novel miRNAs were identified by searching for antisense hits in the reference RNA sequences of the channel catfish. These potential target sequences are involved in immune regulation, transcriptional regulation, metabolism and many other biological functions. The discovery of miRNAs in the channel catfish genome by this study contributes to a better understanding of the role miRNAs play in regulating diverse biological processes in fish and vertebrates.

Yellow catfish (Pelteobagrus fulvidraco) is one of the most important freshwater aquaculture species in China. However, its small size and lower meat yield limit its edible value. Myostatin (MSTN) is a negative regulator of mammalian muscle growth. But, the function of Mstn in fish remains elusive. To explore roles of mstn gene in fish growth and create a strain of yellow catfish with high amount of muscle mass, we performed targeted disruption of mstn in yellow catfish using engineered zinc-finger nucleases (ZFNs). Employing zebrafish embryos as a screening system to identify ZFN activity, we obtained one pair of ZFNs that can edit mstn in yellow catfish genome. Using the ZFNs, we successfully obtained two founders (Founder July29-7 and Founder July29-8) carrying mutated mstn gene in their germ cells. The mutated mstn allele inherited from Founder July29-7 was a null allele (mstnnju6) containing a 4 bp insertion, predicted to encode function null Mstn. The mutated mstn inherited from Founder July29-8 was a complex type of mutation (mstnnju7), predicted to encode a protein lacking two amino acids in the N-terminal secretory signal of Mstn. Totally, we obtained 6 mstnnju6/+ and 14 mstnnju7/+ yellow catfish. To our best knowledge, this is the first endogenous gene knockout in aquaculture fish. Our result will help in understanding the roles of mstn gene in fish.

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