Development of a retroviral construct containing a human mutated dihydrofolate reductase cDNA for hematopoietic stem cell transduction Journal Article
| Authors: | Li, M. X.; Banerjee, D.; Zhao, S. C.; Schweitzer, B. I.; Mineishi, S.; Gilboa, E.; Bertino, J. R. |
| Article Title: | Development of a retroviral construct containing a human mutated dihydrofolate reductase cDNA for hematopoietic stem cell transduction |
| Abstract: | A double-copy Moloney leukemia virus-based retroviral construct containing both the Neo(R) gene and a mutant human dihydrofolate reductase (DHFR) cDNA (Ser31 mutant) was used to transduce NIH 3T3 and mouse bone marrow (BM) progenitor cells. This resulted in increased resistance of these cells to methotrexate (MTX). The transduced BM progenitor cells were returned to lethally irradiated mice. The recipients transplanted with marrow cells infected with the recombinant virus showed protection from lethal MTX toxicity as compared with mock-infected animals. Evidence for integration of the proviral DNA was obtained by amplification of proviral DNA by polymerase chain reaction (PCR) and Southern analysis. Sequencing a portion of the PCR- amplified human DHFR cDNA showed the presence of the mutation. These studies with the human Ser31 mutant DHFR cDNA gave results comparable with those obtained with the mutant murine DHFR cDNA (Leu to Arg22) in developing MTX- resistant BM. The Ser31 mutant human DHFR cDNA is currently being tested for infection of human CD34+ human BM and peripheral blood stem cells in vitro. |
| Keywords: | human cell; mutation; methotrexate; polymerase chain reaction; animal; mice; gene amplification; genetic transduction; transduction, genetic; stem cell; genetic transfection; hematopoietic stem cells; hematopoiesis; dna sequence; dihydrofolate reductase; tetrahydrofolate dehydrogenase; bone marrow transplantation; moloney murine leukemia virus; complementary dna; 3t3 cells; dna, complementary; southern blotting; human; priority journal; article; support, u.s. gov't, p.h.s. |
| Journal Title: | Blood |
| Volume: | 83 |
| Issue: | 11 |
| ISSN: | 0006-4971 |
| Publisher: | American Society of Hematology |
| Date Published: | 1994-06-01 |
| Start Page: | 3403 |
| End Page: | 3408 |
| Language: | English |
| PROVIDER: | scopus |
| PUBMED: | 8193378 |
| DOI: | 10.1182/blood.V83.11.3403.3403 |
| DOI/URL: | |
| Notes: | Export Date: 14 January 2019 -- Article -- Source: Scopus |
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