6-Carboxytetramethylrhodamine (6-TAMRA), with the CAS number 91809-67-5, is a highly popular fluorescent dye in the field of biochemical research. It is a member of the rhodamine family, known for its excellent photostability and high fluorescence quantum yield.
Chemical Formula: C₂₅H₂₂N₂O₅
Molecular Weight: Approximately 430.45 g/mol
Appearance: Usually in the form of a red powder
Solubility: It is soluble in polar organic solvents such as dimethyl sulfoxide (DMSO) and dimethylformamide (DMF), and can also be dissolved in aqueous buffer solutions with appropriate pH values.
1. DNA Sequencing: 6-TAMRA is widely used in fluorescent DNA sequencing. It can be conjugated to primers or nucleotides, and its fluorescence signal can be detected during the DNA sequencing process, allowing for the accurate determination of DNA sequences.
2. Oligonucleotide Labeling: In the field of molecular biology, it is used to label oligonucleotides. This labeled oligonucleotide can be used for various hybridization assays, such as fluorescence in situ hybridization (FISH), which helps in the detection and localization of specific DNA or RNA sequences in cells or tissues.
3. Protein Labeling: It can be used to label proteins to study protein-protein interactions, protein localization, and protein function. The fluorescence properties of 6-TAMRA allow for real - time monitoring of protein behavior in biological systems.
1. Conjugation Reaction: When conjugating 6-TAMRA to biomolecules such as oligonucleotides or proteins, appropriate coupling agents and reaction conditions are required. For example, when conjugating to amino - containing biomolecules, N - hydroxysuccinimide (NHS) esters of 6-TAMRA can be used. The reaction is usually carried out in a buffer solution with a suitable pH (around 7.5 - 8.5) at room temperature or under mild heating conditions for a certain period of time (usually several hours).
2. Purification: After the conjugation reaction, purification is necessary to remove unreacted 6-TAMRA and other impurities. Common purification methods include gel filtration chromatography, high - performance liquid chromatography (HPLC), etc.
3. Fluorescence Detection: Once the labeled biomolecule is obtained, fluorescence detection can be performed. The excitation wavelength of 6-TAMRA is around 555 nm, and the emission wavelength is around 580 nm. Appropriate fluorescence spectrometers or imaging systems can be used to detect the fluorescence signal.
In a recent DNA sequencing project, a research team used 6-TAMRA - labeled primers. They first carried out the conjugation reaction according to the standard protocol. After purification by HPLC, they obtained high - quality labeled primers.
During the DNA sequencing process, they used a capillary electrophoresis system equipped with a fluorescence detector. The 6-TAMRA - labeled fragments produced clear and distinct fluorescence signals, which enabled accurate base - calling. The results showed that the use of 6-TAMRA - labeled primers significantly improved the accuracy and efficiency of DNA sequencing compared to traditional methods. The researchers were able to sequence a large number of DNA fragments in a relatively short time, and the error rate was reduced by about 30%.
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