Green fluorescence protein as a transcriptional reporter for the long terminal repeats of the human immunodeficiency virus type 1

Anindita Kar-Roy, Wei Dong, Nelson Michael, Yen Li*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

12 Scopus citations

Abstract

Using the enhanced green fluorescence protein (EGFP), a transient reporter expression system was established to assess the transcriptional activity of the long terminal repeats (LTR) of primary isolates of the human immunodeficiency virus type 1 (HIV-1). Consistent with the conventional chloramphenicol acetyl transferase (CAT) reporter, EGFP expression, under the direction of HIV-1 LTR, was readily detected in the transient transfection and was elevated by co-transfection of HIV-1 tat-expression vector. Comparing to CAT, however, EGFP expression system has two advantages: (i) Using a fluorescence activated cell sorter (FACS), it was possible to simultaneously measure transfection efficiency and fluorescence intensity of the transfected live cells without the necessity of co-transfection of a reference plasmid for comparing the transcriptional activity of two promoters; and (ii) EGFP expression was readily detected at a DNA concentration where CAT activity was not detectable possibly because the transfectants could be 'gated'. On the other hand, at a higher concentration of DNA, CAT signal became more prominent than that of EGFP, possibly because the enzymatic activity of CAT 'amplified' the signal. EGFP fluorescence detected by FACS was a direct measurement of the expressed chromophore. It is concluded that the system is rapid, reproducible, convenient and useful for quantitative analysis of transcription. Copyright (C) 2000 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)127-138
Number of pages12
JournalJournal of Virological Methods
Volume84
Issue number2
DOIs
StatePublished - Feb 2000
Externally publishedYes

Keywords

  • Green fluorescence protein
  • Human immunodeficiency virus type 1
  • Transcriptional reporter

Fingerprint

Dive into the research topics of 'Green fluorescence protein as a transcriptional reporter for the long terminal repeats of the human immunodeficiency virus type 1'. Together they form a unique fingerprint.

Cite this