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. 2011 Sep 20;152(11):4298–4309. doi: 10.1210/en.2011-1521

Fig. 1.

Fig. 1.

Kiss1-CreGFP targeting construct. A 13.2-kb Afe1-Sal1 fragment, including approximately 5.0-kb DNA 5′of the Kiss1 translation start site and approximately 8 kb on the 3′ side (depicted as broken dark blue arrow with a line through it in the top panel), was subcloned from a C57Bl/6 BAC clone into Bluescript. The targeting construct (depicted in the middle panel) contained a CreGFP fusion protein cassette (red and green arrow inserted in the middle of the first segment of the dark blue broken arrow from the top panel), followed by an frt-flanked SvNeo gene [for positive selection, illustrated by red diamonds (frt sites) surrounding a green arrow (SvNeo)] inserted just upstream of the normal start site for Kiss1. The construct also contained Pgk-DTA and HSV-TK (light blue arrows) genes for negative selection. The frt-Neo gene was removed by breeding the heterozygous mice with FLPer mice. The final construct is depicted in the bottom panel, illustrating a new Kiss1 allele that contains a Cre-eGFP cassette inserted before the native Kiss1 start site.