Systemin is a well - known plant peptide hormone that plays a crucial role in the plant's defense response against herbivores and pathogens. As a leading Systemin supplier, understanding how to detect Systemin in plants is of utmost importance, not only for scientific research but also for ensuring the quality and authenticity of the Systemin products we offer. In this blog, we will explore various methods of detecting Systemin in plants, delving into the scientific principles behind each approach.
1. Immunological Detection Methods
Immunological detection techniques are widely used for detecting Systemin in plants due to their high specificity and sensitivity. These methods rely on the use of antibodies that specifically recognize and bind to Systemin.
Enzyme - Linked Immunosorbent Assay (ELISA)
ELISA is a commonly employed immunological method. It involves coating a microplate with an antibody specific to Systemin. Plant extracts are then added to the wells of the microplate. If Systemin is present in the extract, it will bind to the immobilized antibody. Subsequently, a second antibody conjugated with an enzyme is added. This secondary antibody also binds to Systemin. After washing away any unbound substances, a substrate for the enzyme is added. The enzyme - substrate reaction produces a detectable signal, often a color change, which can be measured photometrically. The intensity of the signal is proportional to the amount of Systemin in the sample.
The advantages of ELISA include its high sensitivity, ability to process multiple samples simultaneously, and relative ease of use. However, it requires the production of high - quality antibodies, and cross - reactivity with other peptides in the plant extract can sometimes be a problem.
Western Blotting
Western blotting is another powerful immunological technique. First, plant proteins are extracted and separated by gel electrophoresis based on their molecular weight. The separated proteins are then transferred onto a membrane. An antibody specific to Systemin is incubated with the membrane. After washing, a secondary antibody conjugated with a detection reagent (such as a fluorescent or chemiluminescent marker) is added. The presence of Systemin is visualized as a distinct band on the membrane.
Western blotting allows for the determination of the molecular weight of Systemin and can provide information about its possible modifications. However, it is more time - consuming and technically demanding compared to ELISA.
2. Mass Spectrometry - Based Detection
Mass spectrometry (MS) is a highly accurate and versatile method for detecting Systemin in plants. It can identify and quantify peptides based on their mass - to - charge ratio (m/z).
Liquid Chromatography - Mass Spectrometry (LC - MS)
In LC - MS, plant extracts are first separated by liquid chromatography. The separated components are then introduced into a mass spectrometer. The mass spectrometer ionizes the molecules and measures their m/z values. By comparing the obtained m/z values with the theoretical m/z of Systemin, the presence of Systemin can be confirmed.
LC - MS offers high resolution and the ability to detect Systemin even in complex plant matrices. It can also provide information about the structure and possible post - translational modifications of Systemin. However, it requires expensive equipment and highly trained personnel.
Tandem Mass Spectrometry (MS/MS)
MS/MS is an extension of mass spectrometry. After the initial mass analysis in the first mass spectrometer, selected ions are fragmented, and the fragments are analyzed in a second mass spectrometer. This allows for the determination of the amino acid sequence of Systemin, which is essential for confirming its identity.
MS/MS is particularly useful when dealing with samples where there may be isobaric or isomeric peptides that could interfere with the identification. It provides detailed structural information about Systemin, but it is also more complex and time - consuming than single - stage mass spectrometry.
3. Molecular Biology - Based Detection
Molecular biology techniques can also be used to detect Systemin at the gene expression level.
Reverse Transcription - Polymerase Chain Reaction (RT - PCR)
RT - PCR is used to detect the mRNA encoding Systemin. First, total RNA is extracted from plant tissues. The mRNA is then reverse - transcribed into complementary DNA (cDNA) using reverse transcriptase. Specific primers for the Systemin gene are used to amplify the cDNA by PCR. The amplified DNA fragments can be visualized by gel electrophoresis.
RT - PCR allows for the detection of Systemin gene expression, which can indicate the physiological state of the plant in terms of its defense response. However, it only provides information about the potential production of Systemin at the transcriptional level and does not directly measure the presence of the peptide itself.
Quantitative Real - Time PCR (qRT - PCR)
qRT - PCR is a more advanced version of RT - PCR. It allows for the quantification of the amount of Systemin mRNA in the sample. During the PCR reaction, a fluorescent dye or a fluorescent - labeled probe is used to monitor the amplification in real - time. The cycle at which the fluorescence signal crosses a threshold (Ct value) is inversely proportional to the amount of the target mRNA.
qRT - PCR provides more accurate quantification of Systemin gene expression compared to traditional RT - PCR. It is useful for studying the regulation of Systemin production in response to different stimuli.
4. Bioassay - Based Detection
Bioassays rely on the biological activity of Systemin to detect its presence. For example, Systemin is known to induce the synthesis of protease inhibitors in plants. A bioassay can be set up by treating plant tissues with a sample suspected to contain Systemin and then measuring the induction of protease inhibitors.
The advantage of bioassays is that they directly measure the biological activity of Systemin. However, they are less specific compared to other methods and can be affected by other factors in the plant extract that may also influence protease inhibitor synthesis.
Conclusion
As a Systemin supplier, we understand the importance of accurate and reliable Systemin detection. Each of the methods described above has its own advantages and limitations. Immunological methods are highly specific and sensitive, mass spectrometry provides detailed structural information, molecular biology techniques offer insights into gene expression, and bioassays measure biological activity. In practice, a combination of these methods may be used to ensure the accurate detection and quantification of Systemin in plants.
If you are interested in purchasing high - quality Systemin for your research or other applications, we invite you to contact us for further discussions. We are committed to providing you with the best products and services. You may also be interested in some of our other peptide products, such as Glycoprotein IIb Fragment (296 - 306), PHM - 27 (human), and Secretin (rat).
References
- Ryan, C. A. (2000). The systemin signaling pathway: differential activation of plant defensive genes. Biochimie, 82(10 - 11), 847 - 853.
- Schilmiller, A. L., & Howe, G. A. (2005). Systemin: a polypeptide signal for plant defensive responses. Current Biology, 15(18), R744 - R747.
- van der Hoorn, R. A., & Jones, J. D. (2004). Proteases in pathogen defense. Current Opinion in Plant Biology, 7(4), 400 - 406.