Abstract
Like other complex retroviruses such as HIV-1, HTLV-1 encodes several regulatory and auxiliary non-structural proteins from overlapping open reading frames through the generation of alternatively spliced mRNAs. HTLV-1 expression is orchestrated by the Tax and Rex regulatory proteins; Tax drives the transcription of the viral genome, while Rex acts at the posttranscriptional level by enhancing the nuclear export and expression of unspliced and incompletely spliced mRNAs. The present chapter is focused on the techniques employed to quantitate HTLV-1 mRNAs in the nuclear and cytoplasmic compartments. To ensure a quantitative transcript-specific detection of the levels of individual HTLV-1 mRNAs in a complex mixture of closely related species, splice junction-specific primers and TaqMan probes were used. As HTLV-1 gene regulation is based on the controlled nucleo-cytoplasmic export of the different viral mRNAs, we quantitated the individual viral transcripts in the nuclear and cytoplasmic fractions.
| Original language | English |
|---|---|
| Title of host publication | Methods in Molecular Biology |
| Publisher | Humana Press Inc. |
| Pages | 325-337 |
| Number of pages | 13 |
| Volume | 1087 |
| ISBN (Print) | 9781627036696 |
| DOIs | |
| Publication status | Published - 2014 |
Publication series
| Name | Methods in Molecular Biology |
|---|---|
| Volume | 1087 |
| ISSN (Print) | 10643745 |
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Keywords
- Alternative splicing
- HTLV-1
- Leukemia
- Nucleo-cytoplasmic fractionation
- qRT-PCR
- Rex
ASJC Scopus subject areas
- Molecular Biology
- Genetics
Cite this
Quantitative analysis of Human T-Lymphotropic Virus Type 1 (HTLV-1) gene expression using nucleo-cytoplasmic fractionation and splice junction-specific real-time RT-PCR (qRT-PCR). / Cavallari, Ilaria; Rende, Francesca; Ciminale, Vincenzo.
Methods in Molecular Biology. Vol. 1087 Humana Press Inc., 2014. p. 325-337 (Methods in Molecular Biology; Vol. 1087).Research output: Chapter in Book/Report/Conference proceeding › Chapter
}
TY - CHAP
T1 - Quantitative analysis of Human T-Lymphotropic Virus Type 1 (HTLV-1) gene expression using nucleo-cytoplasmic fractionation and splice junction-specific real-time RT-PCR (qRT-PCR)
AU - Cavallari, Ilaria
AU - Rende, Francesca
AU - Ciminale, Vincenzo
PY - 2014
Y1 - 2014
N2 - Like other complex retroviruses such as HIV-1, HTLV-1 encodes several regulatory and auxiliary non-structural proteins from overlapping open reading frames through the generation of alternatively spliced mRNAs. HTLV-1 expression is orchestrated by the Tax and Rex regulatory proteins; Tax drives the transcription of the viral genome, while Rex acts at the posttranscriptional level by enhancing the nuclear export and expression of unspliced and incompletely spliced mRNAs. The present chapter is focused on the techniques employed to quantitate HTLV-1 mRNAs in the nuclear and cytoplasmic compartments. To ensure a quantitative transcript-specific detection of the levels of individual HTLV-1 mRNAs in a complex mixture of closely related species, splice junction-specific primers and TaqMan probes were used. As HTLV-1 gene regulation is based on the controlled nucleo-cytoplasmic export of the different viral mRNAs, we quantitated the individual viral transcripts in the nuclear and cytoplasmic fractions.
AB - Like other complex retroviruses such as HIV-1, HTLV-1 encodes several regulatory and auxiliary non-structural proteins from overlapping open reading frames through the generation of alternatively spliced mRNAs. HTLV-1 expression is orchestrated by the Tax and Rex regulatory proteins; Tax drives the transcription of the viral genome, while Rex acts at the posttranscriptional level by enhancing the nuclear export and expression of unspliced and incompletely spliced mRNAs. The present chapter is focused on the techniques employed to quantitate HTLV-1 mRNAs in the nuclear and cytoplasmic compartments. To ensure a quantitative transcript-specific detection of the levels of individual HTLV-1 mRNAs in a complex mixture of closely related species, splice junction-specific primers and TaqMan probes were used. As HTLV-1 gene regulation is based on the controlled nucleo-cytoplasmic export of the different viral mRNAs, we quantitated the individual viral transcripts in the nuclear and cytoplasmic fractions.
KW - Alternative splicing
KW - HTLV-1
KW - Leukemia
KW - Nucleo-cytoplasmic fractionation
KW - qRT-PCR
KW - Rex
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U2 - 10.1007/978-1-62703-670-2_26
DO - 10.1007/978-1-62703-670-2_26
M3 - Chapter
AN - SCOPUS:84934441466
SN - 9781627036696
VL - 1087
T3 - Methods in Molecular Biology
SP - 325
EP - 337
BT - Methods in Molecular Biology
PB - Humana Press Inc.
ER -