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Cat. No.: SRLL-20 (for 20T)

Cat. No.: SRLL-40 (for 40T)

Description

Small RNA 3'-Linker (5'adenylated, 3'blocked) and Ligation Kit is a reagent kit that provides a Universal miRNA Cloning Linker with 5' adenylation and 3' blocking, along with the corresponding ligation enzyme and related reagents. It is used for the ligation of small RNAs, such as miRNAs, which have a 3' hydroxyl group, to an adapter. The small RNA, once ligated to the adapter at the 3' end, can be further used for cloning, high-throughput sequencing library preparation, or PCR detection.

This kit can also be used for the ligation of single-stranded DNA with a 3' hydroxyl group to an adapter, and therefore, it can also be referred to as ssDNA 3'-Linker (5'adenylated, 3'blocked) and Ligation Kit, or 3'-Linker (5'adenylated, 3'blocked) and Ligation Kit for nucleic acids.

The Universal miRNA Cloning Linker provided in this kit has adenylation at the 5' end (also known as 5' pre-adenylation or 5' pre-adenylated) and amino blocking at the 3' end. It is used for the addition of a 3' adapter to RNA molecules with a 3' hydroxyl group. The T4 RNA Ligase 2, truncated provided in this kit recognizes the "activated" 5' adenylated Universal miRNA Cloning Linker and can covalently ligate its 5' end to the 3' hydroxyl group of another single-stranded oligonucleotide without the need for ATP.

When using this kit for miRNA ligation, it effectively prevents self-ligation of miRNAs, ligation between different miRNAs, cyclization of the Linker, or ligation between Linker molecules. Instead, it specifically facilitates the ligation between the 5' end of the Linker and the 3' hydroxyl group of the miRNA.

The sequence of the Universal miRNA Cloning Linker provided in this kit is: 5´-rAppCTGTAGGCACCATCAAT–NH2-3´. This sequence is the same as NEB's S1315S Universal miRNA Cloning Linker.

If this kit is used in a 20µl reaction volume and following the recommended ligation protocol of the kit, it can be used for 20 reactions of Universal miRNA Cloning Linker and ssRNA ligation.

Precautions

• The reagents provided with this kit are prepared using DEPC water and can be directly used for the ligation reaction between single-stranded RNA (with a 3' hydroxyl group) and Universal miRNA Cloning Linker (5' adenylated, 3' blocked).
• When using this kit to ligate ssRNA and Universal miRNA Cloning Linker (5' adenylated, 3' blocked), it is recommended to use a concentration of 10% - 30% PEG8000. Increasing the concentration of PEG8000 appropriately will significantly improve the ligation efficiency.
• This product is intended for scientific research purposes by professionals only. It should not be used for clinical diagnosis or treatment, food or drug purposes, and should not be stored in residential areas.
• For your safety and health, please wear appropriate lab attire and disposable gloves while handling.

Storage

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