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Mediators of Inflammation
Volume 2012, Article ID 495934, 14 pages
Research Article

Altered AKT1 and MAPK1 Gene Expression on Peripheral Blood Mononuclear Cells and Correlation with T-Helper-Transcription Factors in Systemic Lupus Erythematosus Patients

1Department of Cellular Biology and Immunology, Instituto de Parasitología y Biomedicina López-Neyra, (IPBLN-CSIC), Parque Tecnológico Ciencias de la Salud, Avenida Conocimiento s/n, Armilla, 18100 Granada, Spain
2Systemic Autoimmune Diseases Unit, Department of Internal Medicine, San Cecilio University Hospital (SCUH), Avenida Dr. Oloriz, no. 16, Granada 18012, Spain
3Department of Microbiology, University of Alabama at Birmingham, 1720 2nd Ave South, Birmingham, AL 35294, USA
4Department of Dermatology, SCUH, Avenida Dr. Oloriz, No. 16, Granada 18012, Spain

Received 9 May 2012; Revised 21 August 2012; Accepted 3 September 2012

Academic Editor: Eric F. Morand

Copyright © 2012 Sonia Garcia-Rodriguez et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Supplementary Material

qRT-PCR procedures: Reverse transcription (RT) reaction was performed on 1 µg total RNA, 20-µl reaction, containing 200 U SuperScript III reverse transcriptase (Invitrogen), 5 mM random hexamer primers, 0.5mM each deoxyribonucleotide triphosphate (dNTP), 20 U RNase inhibitor, and 1X-RT buffer. Primer sequences and gene-specific primers used are available in Table 2. Gene expressions assays were performed on iQ-Cycler (Bio-Rad), with 3 replicates with iQ-SYBR Green Supermix, 21-microliter reaction (200nM each primer and 10 ng of cDNA). PCR program: 10 min incubation at 95°C, 40 cycles of 15 s at 95°C, and 1 min at 60°C. Specificity of PCR amplification procedure was checked with a heat dissociation protocol (from 65°C to 100°C), after the final cycle of the PCR.

  1. Supplementary Material