The main conclusion of this study is that our novel Split-Cre complementation system introduces both temporal and special control of site-specific recombination using the Cre recombinase enzyme. This system resolved many drawbacks that appeared during the intensive use of Cre recombinase in molecular biology. The complemented protein is almost as effective as Full CRE in recombination activity (~95%). Furthermore, each fragment lacks recombinase activity. This system allows precise genetic manipulation. It has particular importance in neuroscience, because it lacks a selective promoter region to conditionally hit our specific gene on a specific brain tissue. Many websites, such as GenePaint; a digital atlas for the gene expression pattern in the mouse embryo (http://www.genepaint.org/Frameset.html) (35), METscout (http://www.metscout.mpg.de/) (36) and GENESAT A Brain Atlas of Gene Expression (http://www.gensat.org/cre.jsp) has been published. We can determine the appropriate promoter that selectively expresses in the required tissue with the desired expression intensity among these promoters. Recently, the possibility of Cre complementation has been explored by several laboratories. It has advantages over temporal control of Cre recombinase activity. For example, prior genetic marking of cells expressing the first Cre fragment (nCre or cCre), by co-expression of EGFP. Then, the introduction of the second Cre fragment, to obtain the recombinase activity, will allow a direct comparison between cell morphology and dynamic behavior before and after genetic modification (37). Previous trials split the CRE into two fragments such that (19-59, 60-343) (19) and (1-190, 191-343) (20) could restore a maximum of 35% of the full-length CRE recombinase a ..... . middle of article...... L. Spatial and temporal expression of the Cre gene under the control of MMTV-LTR in different lines of transgenic mice. Res transgenic. 2001; 10(6): 545-553.44- O'Neill MT, Phuong T, Healer J, Richard D, Cowman AF. Deletion of Plasmodium falciparum genes using FLP and Cre recombinases: implications for applied site-specific recombination. Int J Parasitol. 2011; 41(1): 117-123.45- Koblizek TI, Siehoff A, Pitt A. Systematic analysis of complex signal transduction pathways using protein fragment complementation assays. Mol Biol Methods. 2013; 986: 179-185.46- Kozak M. Rethinking certain mechanisms invoked to explain translational regulation in eukaryotes. Embarrassed. 2006; 382: 1-11.47- Smith AE, Kalderon D, Roberts BL, Colledge WH, Edge M, Gillett P, Markham A, Paucha E, Richardson WD. The nuclear localization signal. Proc R Soc Lond B Biol Sci. 1985; 226(1242): 43-58.