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. 2014 Dec 2;8:145. doi: 10.3389/fnana.2014.00145

Figure 6.

Figure 6

Validation of nkx2TKD phenotype by independent nkx2.4a TBMO. By designing a second, non-overlapping MO against nkx2.4a (nkx2.4a TBMO) concerns with off-target phenotypes can be addressed. The probability of both nkx2.4a MOs yielding a similar off-target phenotype is significantly lower compared to a single MO only (Eisen and Smith, 2008). The triple knockdown experiments with the nkx2.4a TBMO were analyzed for changes in lhx6 (A,B) and lhx5 (C,D) expression. The observed phenotypes were very similar to the nkx2TKD with the SBMO: the respective expression domains in the pallidum, the posterior tuberculum, the caudal and rostral hypothalamus were depleted in nkx2TKD with the nkx2.4a TBMO, while expression in other anatomical domains, including the alar preoptic region, was not significantly affected. Therefore, the analysis of nkx2TKD with the nkx2.4a TBMO confirmed the results obtained in our study with the nkx2TKD with the nkx2.4a SBMO. Black arrows point at positions of anatomical structures labeled by abbreviation at start of arrow.