Primer binding

Lee R., Kaushik N., Modak M.J., Vi-NAYAK R., Pandey V. N. Polyamide nucleic acid targeted to the primer binding site of the HlV-1 RNA genome blocks in vitro HlV-1 reverse transcription. Biochemistry 1998 37 900-910. 

Kaushik N., Talele T.T., Monel R., Palumbo P., Pandey V.N. Destabilization of tRNA3Lys from the primer-binding site of HIV-1 genome by anti-A loop polyamide nucleotide analog. Nucleic Acids Res. 2001 29 5099-5106. 

Competitive PCR (cPCR) has emerged as the best strategy for controlling the sam-ple-to-sample variability of PCR. In cPCR different templates of similar lengths and with the same primer binding sequences are coamplified in the same tube. This ensures identical thermodynamics and amplification efficiency for both templates. The amount of one of the templates must be known and, after amplification, products of both templates must be distinguishable and separately quantifiable. 

The specificity of bacterial detection by PCR depends on both the degree of homology between the primers and the target DNA, which dictates how well the primers bind to the target, and the annealing temperature at which the target hybridizes to the primers.83 The primers are designed from the... 

Figure 10.3. Mass array, (a) Primer binding (b) primer extension enzyme, ddATP and dCTP/ dGTP/dTTP addition (c) primer terminates (d) primer extension products ready for MALDI-MS (e) MS spectrum of primer extension products. Each addition of a nucleotide to the primer extension product increases the mass by 289 to 329 Da, depending on the nucleotide added. The mass difference is easily resolved by MALDI-TOF, which has the ability to detect differences as small as 3 Da. Printed by kind permission of Sequenom. (See color insert.)...

Initiation of Reverse Transcription and the Primer Binding Site... 

INITIATION OF REVERSE TRANSCRIPTION AND THE PRIMER BINDING SITE... 

Initiation of reverse transcription in HIV-infected cells relies on a critical RNA-RNA interaction between tRNA y s, which is preferentially packaged into the viral particle, and a specific viral RNA seqnence. The 3 -terminaI 18 nucleotides of tRNA y are complementary to the primer binding site (PBS) sequence located in the 5 -Iong terminal repeat (LTR) of the viral RNA genome (Figure 10.3). The UUU anticodon of the tRNA is complementary to and binds to an adenosine rich loop located 8 nucleotides upstream (5 ) of the PBS. This RNA-RNA duplex which is formed when tRNA y s binds to the PBS fits within the active site of HIV-1 reverse transcriptase, bnt mnitiple interactions between the viral RNA and tRNA y are necessary for efficient initiation of reverse transcription. This interaction nucleates the reverse transcription complex which contains viral RNA, reverse transcriptase, tRNA y pl , nncleocapsid p7, and Vpr (Viral protein R), as well as multiple host factors." ... 

The first PCR-based assay for detection of A. niger used sequences from the 18S rRNA gene as the target for primer binding (J imenez etal.,1999). The authors used primers originally published by Melchers et al. (1994),... 

Figure 28-24 Simplified scheme for replication of the RNA genome of a retrovirus. See Sugden.703 PB, Primer-binding site.

B DNA, randomized Region mi DNA, Primer Binding Site Primer E3 DNA, T7 Promotersequence RNA, randomized Region E3 RNA, Primer Binding Site O Matrix Target... 

In almost every published SELEX protocol, the experiment starts with a pool of chemically synthesized ssDNA. The DNA usually consists of a central randomized region of 20-60 nucleotides that is flanked by two constant regions that are necessary for primer binding. 

Primer annealing. The mixture is rapidly cooled to a defined temperature which allows the two primers to bind to the sequences on each of the two strands flanking the target DNA. This annealing temperature is calculated carefully to ensure that the primers bind only to the desired DNA sequences. One primer binds to each strand Fig. 1). The two parental strands do not reanneal with each other because the primers are in large excess over parental DNA. 

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