© 2018, Chartered Inst. of Building Services Engineers. All rights reserved.The biotypes of Culex pipiens complex and Cx. torrentium were molecularly investigated in the mosquito specimens collected from different regions of Kayseri between June and August 2015 in this study. A total of 1052 female mosquito specimens were sampled during the field surveys and 315 (29.9%) out of these were morphologically identified and reserved as Cx. pipiens complex and/or Cx. torrentium. The first step Real time PCR analyses of genomic DNA isolates from these reserved specimens revealed that all the specimens were belong to Culex pipiens complex and there was no Cx. torrentium positivity. Among the isolates determined in Culex pipiens complex, 311 were determined as belong to lineages of Cx. pipiens form pipiens according to second step Real time PCR analyses and there was no positivity for Cx. pipiens form molestus. The remaining 4 isolates showed no amplification for both two biotypes. ACE-2 and CQ11 microsatellite DNA analyses were carried out on the isolates from all specimens in order to confirm the real time PCR results and investigate the hybrid lineages. All the isolates determined as Cx. pipiens form pipiens showed specific banding pattern for pipiens biotype with the related DNA markers in the analyses result in the confirmation of the isolates. The 4 isolates that did not show amplification for both biotypes in Real time PCR assays were designated as the hybrids of Cx. pipiens form pipiens and Cx. pipiens form molestus with the microsatellite DNA analyses. DNA barcoding and phylogenetic analyses were performed on one isolate belong to each Cx. pipiens form pipiens pipiens and hybrids based on mitochondrial cytochrome oxidase I (mt-COI) gene region. Both isolates were determined as new haplotypes according to the sequence analyses and their phylogenetic relations based on haplotypes with the available homolog linages belong to Cx. pipiens complex in GenBank were revealed.