Revista Médica del Hospital General de México (Jul 2024)
Transfection of the H-RAS gene produces genomic instability in the MOLT-4 cell line
Abstract
Objective: The aim of this study was to evaluate the effect of the normal and mutated H-RAS gene on genomic instability and apoptosis in MOLT-4 cells. Material and methods: The MOLT-4 cells were cultured and DNA was extracted. Then, the region that compromises both codons (12 and 13) of the exon 1 in the H-RAS gene was amplified by PCR and sequenced. The pSV2neo, pSV2neo-Raswt, and pSV2neo-Rasmut plasmids were extracted from the DH5-alpha strain, sequenced, and transfected in MOLT-4 cells. Finally, the number of chromosomes was analyzed by means of karyotype and the number of apoptotic bodies was evaluated using acridine orange with propidium iodide. Results: No mutations were found on the H-RAS gene in the region of the 12 and 13 codons of exon 1 in the MOLT-4 cells. On the other hand, the sequences of the pSV2neo-Raswt and pSV2neo-Rasmut plasmids matched with the normal and mutated versions of H-RAS, respectively. An increase in the number of chromosomes per cell with the normal and mutated H-RAS genes was also observed, as well as, the presence of apoptotic bodies. Conclusions: There are no H-RAS gene mutations in the 12 and 13 codons of exon 1 in MOLT-4 cells. The pSV2neo-Raswt and pSV2neo-Rasmut plasmids contain normal and mutated versions of H-RAS, respectively. The normal and mutated H-RAS genes induce genomic instability and apoptosis in MOLT-4 cells. Plasmids pSV2neo-Raswt and pSV2neo-Rasmut contain normal and mutated versions of the H-RAS gene. Normal and mutated H-RAS genes induce genomic instability and apoptosis in MOLT-4 cells.
