Target gene knockdown by 2',4'-BNA/LNA antisense oligonucleotides in zebrafish.
Gene knockdowns using oligonucleotide-based approaches are useful for studying gene function in both in vitro cell culture systems and in vivo animal models. We evaluated the efficacy of 2',4'-bridged nucleic acids (BNA)-modified antisense oligonucleotides (AONs) for gene knockdown in zebrafish. We used the tcf7l1a gene as a model for testing the knockdown efficacy of 2',4'-BNA AONs and examined how the target sites/affinity and RNase H induction activity of 2',4'-BNA AONs affect knockdown efficacy. We found that tcf7l1a gene function was knocked down by 2',4'-BNA AONs that target the start codon and induce RNase H activity. Although nonspecific p53-mediated developmental defects were observed at higher doses, the effective dose of the 2',4'-BNA AONs for tcf7l1a is much lower than that of morpholino oligonucleotides. Our data thus show a potential application for 2',4'-BNA AONs in the downregulation of specific genes in zebrafish.
1 Graduate School of Pharmaceutical Sciences, Chiba University , Chuo-ku, Chiba, Japan .
Motoyuki
M
Itoh
Mizuki
M
Nakaura
Takeshi
T
Imanishi
Satoshi
S
Obika
D000818:Animals; D001483:Base Sequence; D018387:Codon, Initiator; D004625:Embryo, Nonmammalian; D018507:Gene Expression Regulation, Developmental; D055785:Gene Knockdown Techniques; D008969:Molecular Sequence Data; D060172:Morpholinos / Q000235:genetics / Q000378:metabolism; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000235:genetics* / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000138:chemical synthesis / Q000235:genetics* / Q000378:metabolism; D014176:Protein Biosynthesis; D016914:Ribonuclease H / Q000378:metabolism; D058125:Transcription Factor 7-Like 1 Protein / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D015027:Zebrafish; D029961:Zebrafish Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism
D018387:Codon, Initiator; D060172:Morpholinos; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; D058125:Transcription Factor 7-Like 1 Protein; D016159:Tumor Suppressor Protein p53; D029961:Zebrafish Proteins; C477371:locked nucleic acid; C544914:tcf7l1a protein, zebrafish; D016914:Ribonuclease H