Generation of epstein-barr virus immortalized-B lymphocytes cell line of orang asli. Journal of Genetics and Genome Research, 3 (2). ISSN 2378-3648 (2016)
Abstract
Obtaining sustainable source of cells or DNA from a single individual has always been a constraint in biomedical research. Generation of lymphoblastoid cell lines (LCLs) as a surrogate for replacement of isolated peripheral blood mononuclear cells (PBMCs) has substantially accelerated the process of biological investigations. LCLs can be established by transfection of B lymphocytes from peripheral blood with Epstein Barr Virus (EBV), bearing minor genetic and phenotypic alterations. Here we report the generation of LCLs for the indigenous population (Orang Asli) from Peninsular Malaysia. The establishment of cell line is crucial for such isolated populations since the population size is small and sample collections are often challenging. We applied an alternative approach in isolation of PBMCs before proceeding to EBV transfection of Orang Asli samples, of which the BD Vacutainer CPT tubes were used to replace the conventional protocol such as Ficoll-paque or histopaque. We called this approach the "modified method of CPT" (MMC). A total of 59 out of the 64 samples (92%) of Orang Asli were successfully transformed into LCLs by transfecting with EBV supernatant in a single attempt. To verify the success of LCLs transformation and confirm the presence of the human DNA, FCGR3B gene copy number variations of the DNA extracted from the LCLs were typed using Paralogue ratio test-restriction enzyme digest variant ratio (PRT-REDVR) analysis. The genotype results were compared between the DNA obtained from the LCLs and the original PBMCs from the identical individuals, and results revealed 100% concordance rate. The success of the transformation for LCL for the indigenous populations will eventually allow a more comprehensive and advanced biomedical research to be carried out in the next future without the exhaustion of the biomaterials.
| Item Type: | Article |
|---|---|
| Keywords: | Modified method of CPT, Peripheral blood mononuclear cell, Lymphoblastoid cell line, Copy number variation, FCGR3B, Paralogue ratio test-restriction enzyme digest variant ratio (PRT-REDVR) |
| Taxonomy: | By Niche > Genome > Genomes By Niche > Genome > Human Genome Research |
| Local Content Hub: | Niche > Genome |
| Depositing User: | Hazrul Amir Tomyang (Puncak Alam) |
| Date Deposited: | 31 May 2023 09:02 |
| Last Modified: | 31 May 2023 09:02 |
| Related URLs: |
Actions (login required)
 |
View Item |