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Cat. No.: T4RL2T-5k (for 5KU)

Cat. No.: T4RL2T-20k (for 20KU)

Cat. No.: T4RL2T-100k (for 100KU)

Description

T4 RNA Ligase 2, truncated (T4 Rnl2, truncated or Rnl2 (1-249)) is a truncated form of T4 RNA Ligase 2 that contains only the N-terminal 249 amino acids. It can specifically ligate 5'-adenylated DNA or RNA (App-DNA or App-RNA) to the 3' hydroxyl group of RNA.

T4 RNA Ligase 2, truncated has a completely different enzymatic activity compared to T4 RNA Ligase 2. While the latter is an ATP-dependent dsRNA ligase, the former is an ATP-independent single-stranded nucleic acid ligase that relies on 5'-adenylation.

T4 RNA Ligase 2, truncated is commonly used for cloning, high-throughput sequencing library preparation, PCR detection, and other applications involving single-stranded RNA with a 3' hydroxyl group, such as microRNAs (miRNAs). It adds 5'-adenylated DNA or RNA adapters to the 3' hydroxyl end of miRNAs during library construction.

T4 RNA Ligase 2, truncated can only utilize 5'-adenylated single-stranded DNA or RNA as the 3' adapter, which greatly reduces background in ligation reactions. It is often the preferred choice for library preparation of small RNAs.

T4 RNA Ligase 2, truncated does not require ATP for ligation but relies on 5'-adenylated substrates. Even in the presence of ATP, T4 RNA Ligase 2, truncated cannot catalyze the formation of a phosphodiester bond between the 5'-P end of single-stranded DNA or RNA and the 3'-OH end. In contrast, T4 RNA Ligase 1 can catalyze this reaction in the presence of ATP. Therefore, T4 RNA Ligase 2, truncated is particularly suitable for small RNA library preparation, whether for miRNA sequencing or directional mRNA-Seq, as it improves library quality and effectively reduces background in ligation reactions.

Application

T4 RNA Ligase 2, truncated is used for ligating 5'-adenylated single-stranded DNA or RNA to the 3' hydroxyl end of single-stranded RNA. It is also used for connecting 5'-adenylated single-stranded oligonucleotides and small RNAs in cDNA library construction. It is further employed in strand-specific cDNA library construction to connect 5'-adenylated single-stranded oligonucleotides and RNA. In second-generation sequencing, it is utilized for connecting the 3' end of RNA to adapters in miRNA library construction. Additionally, it is used in cDNA library construction for small RNA transcriptome analysis.

Source

Recombinant protein expressed in Escherichia coli, with the expression gene for the N-terminal 1-249 amino acids of T4 bacteriophage T4 RNA ligase 2.

Unit definition

200 units are defined as the amount of enzyme required to achieve 80% ligation of a 31-mer RNA to the pre-adenylated end of a 17-mer DNA (Universal miRNA Cloning Linker) in a total reaction volume of 10 µl in 1 hour at 25°C.

Purity

Free from DNA endonuclease and exonuclease activity, RNAase activity, and protease activity.

Enzyme storage solution

10 mM Tris-HCl (pH 7.5), 100 mM NaCl, 0.1 mM EDTA, 0.1 mM DTT, 50% (v/v) Glycerol.

Inactivation or inhibition

T4 RNA Ligase 2, truncated can be inactivated by heating at 65°C for 20 minutes. Addition of proteinase K or EDTA can also inhibit its activity.

Precautions

• The reagents provided in this product are prepared using water without DNA and RNA enzymes, and can be directly used for the ligation reaction between 3'-hydroxyl single-stranded RNA and 5'-adenylated DNA or RNA.
• Please select the appropriate operating method based on your specific application. Additional reagents may be required, such as RNase inhibitor and DEPC water.
• When using this product to ligate ssRNA and AppDNA, it is recommended to use a concentration of 10%-30% PEG8000. Increasing the concentration of PEG8000 appropriately will significantly improve the ligation efficiency catalyzed by T4 Rnl2, truncated without altering its reaction characteristics.
• This product is for scientific research purposes by professionals only. It is not intended for clinical diagnosis or treatment, and cannot be used for food or drugs. It should not be stored in ordinary residences.
• For your safety and health, please wear appropriate laboratory attire and disposable gloves during operation.

Storage

The minimum shelf life is 1 year at -20°C.

Related:

• T4 RNA Ligase
• T4 RNA Ligase 1 (ssRNA Ligase)
• T4 RNA Ligase 2 (dsRNA Ligase)
• T4 RNA Ligase 2, truncated
• T4 RNA Ligase 2, truncated KQ

SBS Genetech is recognized as one of the global major leading industry players in Ligase Market by third-party market researchers. For more details, please visit Ligase Market - A Global and Regional Analysis Focus on Product, Source, Application, End User, and Country - Analysis and Forecast, 2022-2032