Manuscript Title:

SALIVA BASED ORAL CANCER SCREENING SYSTEM USING GFP EXPRESSING YEAST CELLS

Author:

SIDRA HASNAIN, SANA KHURSHID, Dr. SANA JAVAID AWAN

DOI Number:

DOI:10.17605/OSF.IO/5X83B

Published : 2023-04-10

About the author(s)

1. SIDRA HASNAIN - The University of Lahore, Lahore, Institute of Molecular Biology and Biotechnology 1-Km Defense Road, Lahore, Punjab.
2. SANA KHURSHID - Virtual University of Pakistan, Department of Molecular Biology, Lahore, Pakistan.
3. Dr. SANA JAVAID AWAN - Kinnard College for Women, Lahore, Department of Zoology Lahore, Pakistan.

Full Text : PDF

Abstract

Purpose: Oral cancer poses a great threat to an individual’s life, which can be mitigated effectively if detection and confirmation occur earlier during the disease. Transforming growth factor-β (TGFβ) plays an integral role as a regulator in governing many biological processes. TGFβ level rises in saliva of oral cancer patients. Thus, it could be used as a diagnostic biomarker. This study aimed to develop a reporter system in yeast cells to detect TGFβ as a tool for the non-invasive screening of oral cancer. Methodology: Yeast cells were transformed with transforming growth factor beta receptor II (TGFβRII), and collagen type II alpha chain I (col2a1) (intronic region) containing vector and confirmed through restriction digestion. The recombinant proteins were expressed, optimized and established through gel electrophoresis analysis. Furthermore, recombinant yeast cells were mixed with saliva of oral cancer patients and assessed through enzyme-linked immunosorbent assay (ELISA) and fluorescence microscopy through green fluorescence protein. Results: The significant difference in expression (P value < 0.05) in the expression of TGFβ was found between transformed and non-transformed yeast cells. The presence of TGFβ was confirmed through ELISA in saliva samples of oral cancer patients, while solid-phase sandwich ELISA was performed to verify TGFβ presence in saliva-treated yeast samples. The highest fluorescence was observed in the yeast samples with expression vectors for both Col2a1 and TGFβRII. Conclusion: In the present study, a yeast based green fluorescent protein reporter system was constructed for molecular reporting of early oral cancer through saliva. TGFβRII protein expression in S. cerevisiae was optimized and detected through ELISA and fluorescent microscopy. This co-transformed system can be used as non-invasive diagnosis of oral cancer.


Keywords

Collagen type II alpha 1 chain, Enzyme-linked immunosorbent assay, Gel electrophoresis, Intronic Region, Transformation, Tumor growth factor beta, Tumor growth factor receptor-II.