Reverse transcriptase template

Zhu YY, Machleder EM, Chenchik A, li R, Siebert PD (2001) Reverse transcriptase template switching a SMART approach fiar full-length cDNA library construction. Biotechniques 30(4) 892-897... 

The viruses responsible for AIDS are human immunodeficiency virus 1 and 2 (HIV 1 and HIV 2) Both are retroviruses, meaning that their genetic material is RNA rather than DNA HI Vs require a host cell to reproduce and the hosts m humans are the T4 lymphocytes which are the cells primarily responsible for inducing the immune system to respond when provoked The HIV penetrates the cell wall of a T4 lymphocyte and deposits both its RNA and an enzyme called reverse transcriptase inside There the reverse transcriptase catalyzes the formation of a DNA strand that is complementary to the viral RNA The transcribed DNA then serves as the template from which the host lymphocyte produces copies of the virus which then leave the host to infect other T4 cells In the course of HIV reproduction the ability of the T4 lymphocyte to reproduce Itself IS compromised As the number of T4 cells decrease so does the body s ability to combat infections... 

Section 28 13 HIV which causes AIDS is a retrovirus Its genetic matenal is RNA instead of DNA HIV contains an enzyme called reverse transcriptase that allows Its RNA to serve as a template for DNA synthesis m the host cell... 

Reverse transcriptase j Synthesizes DNA from RNA template. Synthesis of cDNA from mRNA RNA (S end) mapping studies. 

RNA oncogenic vimses have an unusual enzyme, reverse transcriptase, which is capable of making DNA copies from an RNA template. Cells transformed by these retrovimses have been shown to possess DNA transcripts of the viral RNA. It appears that the transformahon from normal to malignant is associated with the acquisition by the cell of viral DNA. 

Reverse transcriptase An enzyme coded by certain RNA vimses which is able to make complementary single-stranded DNA chains from RNA templates and then to convert these DNA chains to doublehelical form. 

After the virus has attached to CD4 and chemokine receptors, another viral glycoprotein (gp41) assists with viral fusion to the cell and internalization of the viral contents. The viral contents include single-stranded RNA, an RNA-dependent DNA polymerase (also known as reverse transcriptase), and other enzymes. Using the single-stranded viral RNA as a template, reverse transcriptase synthesizes a complementary strand of DNA. The single-stranded viral RNA is removed from the newly formed DNA strand by ribonuclease H, and reverse transcriptase completes the synthesis of double-stranded DNA. The... 

This is the enzyme responsible for the synthesis of mitochondrial DNA and the DNA of some viruses, such as adenoviruses. Polymerase -y is very large and consists of a tetramer of identical oligomers, each having a molecular weight of 47,000. Synthetic ribonucleotides are very effective templates in the laboratory, but this mitochondrial enzyme differs from reverse transcriptase in that natural RNAs are poor templates. 

Reverse transcriptase is an enzyme isolated from viruses that contain a genome that is RNA. This viral enzyme makes DNA using RNA as a template. 

Reverse transcriptase. This enzyme is involved in the replication of retroviruses in vivo. It synthesizes a complementary DNA (cDNA) strand using RNA instead of DNA as its template. It is widely used to create a strand of cDNA from mRNA extracted from cells or tissue for cloning or for PCR analysis. 

RNA amplification by PCR has been facilitated by the use of a single heat-stable enzyme. Thus, DNA polymerase from Thermus thermophilus, which has enhanced reverse transcriptase (rT) activity in presence of manganese, can be used with one buffer system. The high temperature used for rT (70°C) to produce a complementary DNA copy from RNA, and the subsequent amplification of DNA at 60°C, increases efficiency by destabilizing secondary structures in the RNA template. This procedure has been used for the amplification of hepatitis C viral RNA (Yl). 

Telomerase is an enzyme in eukaryotes used to maintain the telomeres. It contains a short RNA template complementary to the DNA telomere sequence, as well as telomerase reverse transcriptase activity (hTRT). Telomerase is thus able to replace telomere sequences that would otherwise be lost during replication. Normally telomerase activity is present only in embryonic cells, germ (reproductive) cells, and stem cells, but not in somatic cells. 

Reverse transcriptase is an RNA-dependent DNA polymerase that requires an RNA template to direct the synthesis of new DNA. Retroviruses, most notably HIV, use this enzyme to repHcate their RNA genomes. DNA synthesis by reverse transcriptase in retroviruses can be inhibited by AZT. ddC, and ddl. 

The next stage is to ensure that the recombinant DNA molecule is copied by the enzymes which s)mthesize nucleic acids. These DNA and RNA polymerases synthesize an exact copy of either DNA or RNA from a pre-existing molecule. In this way the DNA polymerase duplicates the chromosome before each cell division such that each daughter cell will have a complete set of genetic instructions which are then passed to the newly formed RNA by RNA polymerase. While both DNA and RNA polymerase require a preformed DNA template, some viruses (such as HIV) have an RNA genome. To duplicate that genome, and incorporate it into a bacterial or mammalian cell, the viruses encode a reverse transcriptase enzyme which produces a DNA copy from an RNA template. 

DNA-directed DNA polymerases [EC 2.7.7.7], also called DNA nucleotidyltransferases (DNA-directed), are enzymes that catalyze the DNA template-directed extension of the 3 -end of a nucleic acid strand one nucleotide at a time. Thus, n deoxynucleoside triphosphates produce n pyrophosphate (or, diphosphate) ions and DNA . This enzyme cannot initiate the synthesis of a polymeric chain de novo it requires a primer which may be DNA or RNA. RNA-directed DNA polymerases [EC 2.7.7.49], also referred to as reverse transcriptases, DNA nucleotidyltransferases (RNA-directed), and revertases, are enzymes that catalyze the RNA template-directed extension of the 3 -end of a nucleic acid strand one nucleotide at a time. Thus, n deoxynucleoside triphosphates produce n pyrophosphate (or, diphosphate) ions and DNA . As was the case above, this enzyme cannot initiate the synthesis of a polymeric chain de novo it requires a primer which may be DNA or RNA. 

RNA to initiate cDNA synthesis. All cellular mRNA contains multiple repeats of adenine bases (poly-A tails). Therefore the complementary thymine bases (oligo-dT) can be used as a primer that binds to the mRNA template required for the reverse transcriptase to synthesize the cDNA. In the case of pancreatic mRNAs (Figure 4.2), the signihcantly higher mRNA for insulin compared with other proteins allowed success in isolating the insulin-specihc cDNA. Subsequent insertion of cDNA into a bacterial expression vector allowed the production of functional insulin that is now marketed as a successful therapeutic product (Figure 4.2). 

The retroviral genomic RNA serves as the template for synthesis of a double-stranded DNA copy, the provirus (Figure 49-4). Synthesis of the provirus is mediated by a virus-encoded RNA-dependent DNA polymerase, or reverse transcriptase. The provirus is translocated to the nucleus and is integrated into host DNA. Transcription of this... 

A third alternative starts with an extract of RNA, not DNA. Mature eukaryotic mRNA contains a long run or tail of adenine residues at its 3 end. The poly(rA) tail can be hybridized with an oligomer of thymine residues, and the oligo(dT) can then be used as a primer for a particular kind of DNA polymerase known as reverse transcriptase. This enzyme, a polymerase associated with retroviruses, will use RNA as a template to make a complementary DNA copy of the RNA, creating a DNA-RNA double-stranded hybrid. In another round of synthesis, the enzyme can replace the RNA strand entirely with DNA, so that the RNA-DNA hybrid is completely converted to double-stranded DNA containing an exact copy of the original RNA sequence. This DNA molecule is known as cDNA because it has a strand that is complementary to (or a copy of) the original RNA. 

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