Figure 2

Procedures of experiments testing transposon activity in Ciona. (a) Excision assay. A mixture of transposon vector and in vitro transcribed transposase mRNA was injected into Ciona one-cell embryos. After these embryos reach the late tailbud stage, DNA was extracted from embryos and was subjected to polymerase chain reaction analyses to identify the excision. (b, c) An example of excision assay: a Minos vector (pMiFr3dTPOG-B1PM) with an insert longer than 12 kilobases was examined. Excised bands are detected only when the transposase mRNA is co-injected. (d) Interplasmid transposition assay. A donor vector containing a Minos construct and a target vector are introduced into Ciona embryos with transposase mRNA. Plasmids are recovered from Ciona embryos and are used to transform Escherichia coli. The occurrence of transposition was monitored by selection of E. coli using antibiotics. bp, base pairs; ORF, open reading frame.