3. SweScript RT II (second generation) First Strand cDNA Synthesis Kit, 50 rxns $420,

SKU: G3332-50
$420.00
In Stock

Description

Product Information

Product Name Cat.No. Spec.
SweScript RT II First Strand cDNA Synthesis Kit G3332-50 50 rxns
G3332-100 100 rxns

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

Component Number Component G3332-50 G3332-100
G3332-1 SweScrip RT II Enzyme Mixa 50 μL 100 μL
G3332-2 5×Reaction Bufferb 200 μL 400 μL
G3332-3 Oligo (dT)18 Primer (100 μM) 50 μL 100 μL
G3332-4 Random Hexamer Primer (100 μM) 50 μL 100 μL
G3332-5 Nuclease-Free Water 1 mL 1 mL
Product Manual

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);

Component Volume
5×Reaction Buffer 4 μL
Oligo (dT)18 Primer (100 μM) 1 μL
or Random Hexamer Primer (100 μM) or 1 μL
or Gene Specific Primer (2 μM) or 1 μL
SweScrip RT II Enzyme Mix 1 μL
Total RNA/mRNA 0.1 ng-5 μg / 10 pg-0.5 μg
Nuclease-Free Water Add to 20 μL

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:

Temperature Time
25℃a 5 min
55℃b 5-15 min
85℃ 5 s

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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