The (RT-PCR), which is one of many embodiments of the polymerase chain reaction (PCR) Reverse transcription-polymerase chain reaction. This technique is used in the field of molecular biology for detecting the expression levels of RNA widely. RT-PCR has been confused real-time polymerase chain reaction and scholars and students (QPCR) often. However, they are different techniques clearly. By creating a (cDNA) the additional DNA from RNA transcription, RT-PCR, are used for the qualitative detection of gene expression but, QPCR was quantified amplification of DNA using the fluorescent probe . Quantitative PCR, quantitative real-time PCR, [3], QPCR also serves as a real-time quantitative PCR. Conventional PCR and RT-PCR, by amplifying the application of the two techniques in this way, to generate multiple copies of DNA isolation of specific, but is fundamentally different. Conventional PCR is used merely for the exponential amplification of DNA sequences. RT-PCR was used to using reverse transcriptase, cloning a gene that is expressed by reverse transcription of RNA of interest in the complement of the DNA. The cDNA then was newly synthesized was amplified using conventional PCR.

In addition to the qualitative study of gene expression by including an equivalent QPCR, RT-PCR may be used to quantify the RNA absolutely and relatively. Often, combined technique described by (real-time quantitative RT-PCR sometimes) Quantitative RT-PCR, real time RT-PCR is an abbreviated RRT-PCR QRT-PCR, or RT-QPCR,. Most powerful compared to other methods for quantitative RNA by Northern blotting, detecting the level of the QRT-PCR RNA, can be considered quantitative analysis with high sensitivity. Infection and expression of one or more genes and this is used to analyze the expression patterns for the identification of disease often.

Quantification of mRNA using RT-PCR may be achieved either two-step reaction or one step. Difference between the two approaches is the number of tubes used in the course of the procedure. The approach of one stage, all of the cDNA synthesis reaction by PCR amplification occurs in the tube. On the other hand, require a reverse transcriptase reaction, the two-stage reaction was performed in a separate tube PCR amplification. The one-step method, I thought to minimize the deviation from the experiment of the enzyme reaction of all in the environment. However, it is susceptible to degradation in the approach of a single step, and the use of this approach, starting template RNA does not recommend that you repeated analysis of the same sample. Furthermore, the one-step approach, it is less accurate than a two-step approach was reported. The main advantage of the two-step approach, analysis with high reproducibility shows a correlation coefficient in the range of 0988-0974. Removal of primer dimers, so can be achieved by simple modification of the melting temperature, for example, be used DNA binding dye SYBR Green or the like, it is the preferred method of analysis. A disadvantage of two-stage approach is sensitive to contamination by more frequent handling samples.

Serum prostate-specific antigen (PSA) assay has a high false positive rate. Marker PSA is to provide diagnostic information is limited. As well as tumor, multi-marker test for detection of potentially fatal, provides a clinical need that is not met. Using the nanoString nCounter system, 20 – as a means for detecting prostate cancer, gene expression of the gene multiple test was developed based on the digital count gene RNA transcripts urine. In this test, centrifuged to pellet void urine, cells were amplified preselected hybridization probe sets the RNA was purified. Measured by DNA microarray, amplification test cell line RNA is not a gene rich level and the number of matches and bias considerable, but will be displayed. For data analysis, each number, to compare β2 microglobulin, and gene family. I have analyzed urine samples of five preoperative patients, and 2 non-cancer. Pathology information is restored. The low June 06 and Gleason scores and prostate cancer markers noncancerous low, are known, the signal for the majority of genes, were positive when. In contrast, one instance Gleason 4 Figure 5 is a strong signal for all associated with cancer markers including CD24. The non-cancer, there is shown a signal of cancer marker of 6 all, there is a possibility that undiagnosed, harbor cancer this person. Analyze the natural wastes, this multiplex assay can be used to stratify patients and early detection potential screening of cancer. The diagnostic information, I got from RNA signature associated with the cells of prostate cancer.

The transfer, simple presence of tRNA molecule begins full disassembly eventually. During his life, one molecule of the mRNA may be edited processed and transported to previous translations. While prokaryotic molecules often, mRNA molecules of eukaryotes requires the transport and processing a wide range of. RNA is when it is transcribed from DNA. If necessary, the transfer time, DNA polymerase makes a copy of the DNA of gene mRNA. This process is similar in eukaryotes and prokaryotes. The essential difference is that it so that after the start of transcription is performed promptly processed to binding RNA polymerase in prokaryotes, the mRNA processing enzymes during transfer.

, In the processed products, the pre-mRNA or precursor mRNA, it is called the completely processed at a time, it is called mature mRNA or partially untreated short. processing of mRNA varies widely between archaea eukaryotes, and bacteria. MRNA in non-eukaryotic organisms are mature in nature of the transfer, and does not require treatment With the exception of rare cases. However, eukaryotic mRNA precursors, requires a large-scale processing. I is a modified nucleotide guanine that have been added to the 5 or “front” – (referred to as RNA m7G cap or methyl guanosine cap RNA cap, 7 RNA also) cap ‘end of the messenger RNA of eukaryotic transcription initiation soon after 5 ‘. 5 ‘cap 7 are linked via a connection and the transfer nucleotide 5′-5’-triphosphate, – the first is constituted by methylguanosine residue terminal. Her presence is important for recognition by protection from RNase and the ribosome.

Further cap attached to the transfer, the transfer, since each interact and displayed together. After the start of transcription, the 5 ‘end of the mRNA, download is synthesized attached to the cap synthetic complex associated with the RNA polymerase. This enzyme catalyzes the complex chemical reactions which are required for sealing the mRNA. Synthesis proceeds as biochemical reaction of multi-stage. Splitting is a process which is characterized in that it is modified to remove the areas that remain, including specific region of the non-coding sequences called introns, protein coding sequences called exons pre-mRNA. Sometimes, allows gene encodes multiple proteins, pre-mRNA message may be a spliced ​​many different ways. This process is called alternative splicing. Normally, drag is called spliceosome performed by RNA-protein complex but, RNA molecules, some are able to catalyze the splicing of its own.

Polyadenylation, and is connected to the part covalent bond polyadenylyl messenger RNA molecule. In eukaryotes, RNA of all but the histone molecules (of mRNA) was polyadenylated at the 3 ‘end. Poly protein (A) tail to help protect the mRNA from degradation by exonucleases. Also for the termination of transcription, polyadenylation is an important export of mRNA from the translation of the core. mRNA can in order to not interfere with the exonuclease degradation (A) tail act easily and poly now be polyadenylated in prokaryotes.

Immediately after transcription and polyadenylation of the RNA from the DNA. Transfer is complete, the chain mRNA is cleaved by the action of the endonuclease complex associated with the RNA polymerase. After the mRNA is cut, about 250 adenosine residues are added to the end of the 3 ‘cleavage site of the free. This reaction is catalyzed by a polymerase polyadenylation. As alternative splicing, and may be one or more embodiments of the polyadenylated mRNA. Polyadenylation site of mutation. The primary RNA transcript of a gene, cut with a poly A addition site, 100 to 200, is added to the 3 ‘end of the RNA. In this site, if you have changed the result of the stable mRNA and unusually long. Beta globin several mutations to change this site: AATAAA is an example -> AACAAA. Moderate anemia is the result.