SweScript RT II First Strand cDNA Synthesis Kit

Product Information

Description

This product uses modified mutant reverse transcriptase to synthesize first-strand cDNA with high efficiency using total RNA or mRNA as template. The kit contains all the reagents needed to synthesize high-quality first-strand cDNA, and provides two kinds of primers for cDNA synthesis: Random Hexamer Primer and Oligo (dT)18 Primer, the single-stranded cDNA product can be directly used for subsequent PCR or qPCR reactions. SweScript RT II (Reverse Transcriptase) is a mutant Reverse Transcriptase obtained through in vitro evolutionary screening based on M-MLV Reverse Transcriptase. SweScript RT II has no RNase H activity. It avoids the degradation of RNA in the DNA/RNA hybrid template during the synthesis of first-strand cDNA, so as to ensure the synthesis amount and length of first-strand cDNA. Compared with the wild-type enzyme and the first-generation SweScript RT I, the thermal stability and cDNA synthesis efficiency of the second-generation SweScript RT II were further improved. It could efficiently synthesize cDNA in the range of 42-65, and the reverse transcription reaction could be completed in 5 min at the earliest, which was especially suitable for RNA reverse transcription with complex structure.

Storage and Handling Conditions

Wet ice transportation；Store at -20 for 12 months.

Component

a：include RNase Inhibitor；

b：include dNTP Mixture and Mg²⁺.

Assay Protocol

First strand cDNA synthesis step

1. Preparation of reverse transcription reaction system (20 μL reaction system is recommended)；

Note: For high GC or complex templates, RNA templates, reverse transcription primers, nuclease-free water can be pre-mixed and held at 65 ° C for 5 min, then quickly cooled on ice. And then you add the other components.

2. Mix gently and centrifuge；

3. Reverse transcription program Settings：

a：If Random Hexamer Primer is used, the subsequent reaction is carried out after incubation at 25 for 5 min. If Oligo (dT)18 Primer or Gene Specific Primer is used, the reaction can be directly carried out at 55；

b: For high GC or complex templates, the reverse transcription temperature can be increased to 65 ° C.

Note:

1. Wear disposable gloves during operation to avoid RNase contamination.

2. Reverse transcription products can be stored at -20 for a short time. If long-term storage is required, it is recommended to store at -80 after packaging to avoid repeated freezing and thawing.

3. If the template is of eukaryotic origin, it is recommended to select Oligo (dT)18 Primer, which is paired with the 3' Poly A tail of eukaryotic mRNA to obtain the highest yield of full-length cDNA.

4. Random Hexamer Primer or Gene Specific Primer should be used for reverse transcription of prokaryotic RNA.

5. Random Hexamer Primer is widely applicable to mRNA, rRNA, tRNA, small RNA, LncRNA and other templates.

6. If reverse transcription is followed by qPCR experiment, Oligo (dT)18 Primer can be mixed with Random Hexamer Primer to make the cDNA synthesis efficiency of each region of mRNA identical, which is helpful to improve the authenticity and repeatability of quantitative results. 

For Research Use Only!

Ver. No.: V1.0-202111

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