Best Answer

To understand the answer to this question, you have to understand what quantitative real time PCR (qRT-PCR) is measuring. Typically qRT-PCR is used to determine changes in gene expression at the mRNA level between different samples. First RNA is extracted from a sample, and reverse transcriptase is used to convert all the RNA to DNA, which is termed complimentary DNA or cDNA. If you start out with a certain amount of mRNA in each sample, the ensuing cDNA will be proportional to it, and will now be compatible with the polymerase chain reaction.

Once you have cDNA, using DNA primers specific to the gene you want to measure, you amplify your gene of interest. Assuming 100% efficiency, each cycle will double the amount of cDNA from the previous cycle. A qRT-PCR machine measures the relative level of cDNA after each cycle by reading the fluorescence from a probe that binds cDNA, and only fluoresces when bound. (There are also other probes but that's another discussion.)

At the end of the reaction, you will have exponential J shaped curves corresponding to each sample that flatten out at the top. If one sample had less of the gene you are measuring, its curve will be farther to the right, indicating that it took longer for the cDNA to amplify to any level (or fluorescence value) on the Y axis. If you draw a horizontal line through the curves in their exponential region, and then solve for the cycle number it took to get to the point where the curve and your line intersect, you will get the crossing point.

The significance of this is that now you have some value by which to compare samples with. If one samples crossing point is 20 and another's is 22, the sample with a crossing point of 20 had 4 times as much cDNA to begin with of the target gene (2^2), or generally 2^dcp (difference in crossing points) gives the relative difference in starting cDNA.

User Avatar

Wiki User

10y ago
This answer is:
User Avatar

Add your answer:

Earn +20 pts
Q: What is the crossing point in real time PCR?
Write your answer...
Still have questions?
magnify glass
Related questions

Differentiate between quantitative and real time PCR?

: Differentiate between quantitative and real time PCR.

HBV Quantitative real time PCR blood?


What is the defference between Real-time PCR and reverse transcriptase PCR?

Difference between real time PCR and reverse transcription PCR is as follows:- 1. Real time PCR is donated as qPCR and on the other hand reverse transcription PCR is denoted as RT-PCR. 2. In qPCR, the template used is single strand DNA strand whereas in the RT-PCR, the template used in process is single strand of RNA. 3. The real time PCR enables both quantification as well as detection of the DNA in the real time whereas the RT-PCR enables only the quantification of the RNA and it is little bit slower process then the qPCR as it first produce the cDNA from the template RNA strand and then process it in the similar fashion as the traditional PCR.

How long has real time pcr been around?

Real time PCR has been around for over twenty five years. It was revolutionized in the mid 1980's. Real time PCR helps view how much is increased in a DNA sample.

What is a Taqman Real Time PCR?

One can use a PCR to amplify and quantify a certain DNA molecule. A TaqMan real-time PCR is a certain type of PCR which uses the TaqMan method to increase its specificity.

What is a simple explanation of real time pcr?

Real time pcr is a method for making decisions based on real time facts that can be measured in quantity. It is very scientific and not easy to explain simply.

Where can one find an on-line tutorial for Real Time PCR?

One can find an on-line tutorial for Real Time PCR on the official for the University of South Carolina School of Medicine. The website provides an article written by Dr. Margaret Hunt detailing Real Time PCR.

What are the rt pcr?

Real time PCR is uded to amplify and simultaneously quantify a targeted DNA molecule.

What are the different types of polymerase chain reaction techniques?

types of pcr: AFLP -PCR. Allele-specific PCR. Alu-PCR. Assembly -PCR. Assemetric -PCR. Colony -PCR. Helicase dependent amplification. Hot start pCR. Inverse -PCR. Insitu -pCR. ISSR-PCR. RT-PCR(REVERSE TARNSCRIPTASE). REAL TIME -PCR

What are the differences between conventional pcr andreal time pcr?

PCR allows amplification of DNA for a specific gene, after too many cycles of PCR the result will reach saturation, basically meaning all of the DNA has been amplified. Conventional PCR will basically tell you whether or not a gene is expressed in your sample. This can be done semi-quantitavely if the PCR is performed for a low number of cycles, ie it will tell you whether one sample expresses more of your gene of interest than another sample. The results are seen by separating the PCR products by agarose gel/ethidium bromide electrophoresis. Real-time PCR will record exactly what cycle of PCR a detectable level of amplified product became detectable, giving a far more accurately quantifiable estimation of gene expression.

When was Real-time PCR first performed?

The first real-time PCR was first performed in 1993. By 2009, there were seven manufactories that has made as many as eighteen different models. Its a machine that amplifies DNA.

Where online can one learn about real time PCR?

If one were looking to learn more about real time PCR, one should start with Invitrogen. Invitrogen includes learning material which includes webinars and videos.