We then deter mined the amount of HIV 1 DNA by quantitative real

We then deter mined the amount of HIV 1 DNA by quantitative real time PCR. The amount of strong stop cDNA from the cytoplasm of nevirapine treated cells due to natural ERT was subtracted so that only DNA synthesized within the infected cells was measured. We found an approximately twofold lower count of the strong stop DNA in the cells infected enzyme inhibitor with NL 1084 recombinants. We do not believe Inhibitors,Modulators,Libraries that this difference is due to the ability of Inhibitors,Modulators,Libraries nevrapine to inhibit subtype B and C RT differently, because it has been shown that in vitro 10 uM nevira pine inhibited wild type RTs from both subtype B and C viruses by over 100 fold. Analysis of the cDNA accumulation in Sup T1 cells infected with recombinant viruses carrying C terminal Gag products, protease, and RT polymerase domains from different subtype C isolates displayed a significantly decreased level of both early and late reverse transcription products at 24 h post infection.

This result shows the similar effect of the Inhibitors,Modulators,Libraries Pol fragment containing RT polymerase domain from three different isolates of subtype C virus Inhibitors,Modulators,Libraries on the reverse transcription, in spite of individual polymorphism of the AA sequences of RT and different dynamics in disease progression in patients infected with these viruses. Our findings suggest that observed differences in reverse transcription efficiency are dependent on the viral subtype. Since RTCs are undergoing proteasome mediated degradation in the cytoplasm and two thirds of them have been shown to be degraded by several hours post infection, the ratio of the reverse transcription pro ducts in cells infected with different virus strains shown in previous experiments, could be affected by intracyto plasmic degradation of RTCs.

To minimize the effect of host cell mediated degradation of RTCs on reverse transcription, we quantitatively analyzed the cDNA in RTCs isolated from the cytoplasm during the first five hours after infection with subtype B NL4 3, subtype C 1084i, or with chimeric viruses NL polL and 1084 polL. Since Inhibitors,Modulators,Libraries NL and 1084 viral vectors have different tropism, all viruses were pseudotyped with Env glycoprotein of the amphotropic murine leukemia virus. To ensure similar levels of Navitoclax order viruses have entered regardless of the virus backbone and source of the inserted fragment, we measured p24CA content in the RTCs isolated at 1 h after infection, since capsid protein was shown to remain associated with the viral core for hours after infection until completion of the reverse transcription. We found that the p24CA level was similar in early RTCs within virus strains of the same backbone. Differences in p24CA levels between control backbone and chimeric viruses did not exceed 20%.

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