Phi(ɸ)Script™ 1st strand cDNA synthesis contains all the individual components which are required for first strand cDNA synthesis. It contains DX/DT PhiScript reverse transcriptase. It’s an engineered form of Moloney Murine Lukemia Virus reverse transcriptase (M-MuLV- RT). Point mutations are introduced in ɸScript reverse transcriptase so as to reduce RNAse H Activity and to improve its thermal stability. Good cDNA yields can be obtained when reverse transcribed at 42-60°C.
RNA upto 2µg is supported per reaction.
High capacity cDNA synthesis with stable yield.
Efficient reverse transcription upto 60℃.
All the ingredients are given in separate vials for user flexibility.
Contains incredible PhiScript reverse transcriptase.
High tolerance to RNAses.
Accurate results. Simple. Incredible prices.
Ultrafast delivery across India. Less than 3 hrs for Namma Bengaluru.
The Phiscript cDNA Kit is a comprehensive solution tailored for efficient and reliable cDNA synthesis. Engineered to meet the demands of diverse research applications, this kit combines a range of essential components to streamline the process of converting RNA into complementary DNA (cDNA).
At its core, the kit features flagship Phiscript reverse transcriptase, a high-performance enzyme renowned for its robust and accurate reverse transcription capabilities. This enzyme is complemented by a meticulously formulated 5X reaction buffer, optimizing reaction conditions for maximum efficiency and yield.
To ensure versatility and adaptability, the Phiscript cDNA Kit includes a mix of random hexamers and oligo(dT) primers, catering to different RNA species and offering flexibility in target selection. Additionally, the presence of dithiothreitol (DTT) helps maintain enzyme stability and activity throughout the reaction, enhancing overall performance.
Furthermore, the kit incorporates a RNase inhibitor enzyme, safeguarding RNA integrity and preventing degradation during the reverse transcription process. Coupled with a dNTP mix and nuclease-free enzymes, researchers can trust in the purity and quality of their synthesized cDNA, free from contaminating nucleases.
Whether used for gene expression analysis, molecular cloning, or other downstream applications, the Phiscript cDNA Kit provides researchers with a reliable, efficient, and user-friendly solution for cDNA synthesis, empowering them to unlock new insights and discoveries in their fields of study.
DX/DT cDNA synthesis kit can be used for cDNA library preparation and also for gene specific assays. Users has the flexibility to choose reagents of their choice (Hexamers/OligodT/Primers) during reverse transcription. The kit also contains Ribonuclease inhibitor which is a crucial element to protect template RNA from RNAse. Make sure inhibitor enzyme is added upfront before RNA template is added to the reaction vial.
Details
Total human RNA was isolated from skeletal cells. Exon junction primers were designed for human integrin mRNA transcript.
cDNA Synthesis conditions
RNase Inhibitor: 1µL
Random Hexamers: 1µL
OligodT Primers: 1µL
dNTP Mix: 1µL
5X RT Reaction buffer: 4µL
100mM DTT: 1µL
PhiScript™ Reverse Transcriptase: 1µL
RNA template: 1µL (Upto 1µg)
Nuclease Free Water to make up till 20µL.
cDNA synthesis
25℃ (10 minutes)
42℃ (30minutes to 60 minutes)– Reverse Transcription
85℃ (10min) — Inactivates RT
-20℃ — Storage
PCR Conditions
cDNA from the condition was tested and mastermix ability was checked for 3 different sizes of the same human integrin transcript ( 150bp, 650bp and 1.6kb). Appropriate forwad and reverse primers were used.
General PCR conditions:
cDNA: 2µL, For/Rev primers: 1µL each, DX/DT Extremo mastermix: 25µL and make up 50µL using nuclease free water.
PCR Cycles:
95℃ – 2min, [95℃-30s, 60℃-30s, 72℃-1min/kb]30cycles, Final extension, 72℃ for 2min.
5µL of the sample was loaded on 1.5% agarose gel and run for 30 minutes.
DX/DT cDNA synthesis kit can be used for gene specific assays where cDNA of specific gene of interest is made instead of whole library. You have to use only ‘reverse primer’ specific to gene of interest in cDNA synthesis step. Random Hexamers/oligodT shouldn’t be used.
Details
Total human RNA was isolated from skeletal cells. Exon junction primers were designed for human integrin mRNA transcript.
cDNA Synthesis conditions
RNase Inhibitor: 1µL
Reverse Primer (10µM): 2µL
dNTP Mix(10mM): 1µL
5X RT Reaction buffer: 4µL
100mM DTT: 1µL
PhiScript™ Reverse Transcriptase: 1µL
RNA template: 1µL (Upto 1µg)
Nuclease Free Water to make up till 20µL.
cDNA synthesis
42-55℃ (30minutes to 60 minutes)– Reverse Transcription using only ‘reverse primer‘
85℃ (10min) — Inactivates RT
-20℃ — Storage
PCR Conditions
cDNA from the condition was tested and mastermix ability was checked for 1.6kb. Appropriate forwad and same reverse primers were used.
General PCR conditions:
cDNA: 2µL, For/Rev primers: 1µL each, DX/DT Extremo mastermix: 25µL and make up 50µL using nuclease free water.
PCR Cycles:
95℃ – 2min, [95℃-30s, 60℃-30s, 72℃-1min/kb]30cycles, Final extension, 72℃ for 2min.
5µL of the sample was loaded on 1.5% agarose gel and run for 30 minutes.
Long time storage: -20℃, 2 year.
Protocol for this product can be found here.
All the products are for research purpose only. Not for human diagnostics.
