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High Efficient Reverse Transcription Kit

Since first-strand cDNA synthesis is less than 100% efficient, I doubt whether it can make the smear many times brighter than the mRNA in the original total RNA.)

The intensity of the smear depends on how much you load.

I assume that the more cDNA strands that can be synthesized from one mRNA strand during the incubation step, the brighter the resulting smear on the gel.)

Good question.

High Efficient Reverse Transcription Kit for Real-time PCR

High Efficient cDNA synthesis kit with gDNA remover

Cultured cell lysis & cDNA Synthesis kit for real-time PCR

I assume that the more cDNA strands that can be synthesized from one mRNA strand during the incubation step, the brighter the resulting smear on the gel.)


Any ideas would be much appreciated!

Anton the image will depend on the type of primer used in the synthesis: Random hexamers will copy any RNA including ribosomal RNA, while oligo dTs will only copy poly-A-tailed mRNA. 1.

Strangely, I also don't know of any first-strand cDNA synthesis kits that provide such photos in their manuals.

I think most people don't show it because they don't bother to check it, and it's generally not of high scientific interest.

CDNA Synthesis Kits | Life Science Research | Bio-Rad

(I assume that the rRNA bands will be relatively less bright when using oligo-dT primers than when using the random primers, since the latter also creates cDNA from the rRNA.)

3) If I get a smear without any rRNA bands (sometimes a rather bright smear), is this fine, or does it mean that the cDNA synthesis didn't work very well and that the smear is the result of RNA breakdown?

4) On a more in-depth technical point: can M-MuLV or AMV reverse transcriptase re-initialize cDNA synthesis many times over from the same mRNA template strand?

cDNA synthesis kits

ReverTra Ace-α-® is an efficient and convenient kit to synthesize high quality cDNA. This kit contains the highly efficient reverse transcriptase "ReverTra Ace®", as well as other components optimized for the synthesis of long cDNAs suitable for RT-PCR. ReverTra Ace® is an M-MLV reverse transcriptase that has been improved by point mutation technology. ReverTra Ace® has two mutations in the polymerase and RNase H domains.

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cDNA Synthesis Kits - SMART cDNA Synthesis


Other cDNA synthesis kits for endpoint PCR applications.

When synthesizing a protein, DNA is transcribed into mRNA which is then translated into a protein. One difference between eukaryotic and prokaryotic genes is that eukaryotic genes often contain introns which are not coding sequences, in contrast with exons, which are DNA coding sequences. During transcription, intronic RNA is excised from the RNA primary transcript and the remaining pieces of the RNA primary transcript are spliced back together resulting in processed mRNA. The mRNA code is then translated into an amino acid chain that comprises the newly made protein.

Using reverse transcriptase polymerases, DNA can be synthesised from mRNA and total RNA. Thus it is a 'complementary' copy of the mRNA, and is thus called complementary DNA (cDNA). cDNA forms the substrate for the majority of qPCR gene expression experiments.

Toyobo Life Science Department -Products cDNA synthesis kits-

First strand cDNAs synthesis by reverse transcription the products are single stranded. some buffer do not contain DTT, in this case add DTT to 10 mM final. Dec 15, 2006. polymerase RdRp for cDNA synthesis. Dithiothreitol used in the cDNA synthesis step was found to significantly decrease the detection.

cDNA Synthesis Kits Kits on ZAGENO

This kit includes master mix reagents for reverse transcription and for the removal of genomic DNA [DNase I treatment]. In many cases, total RNA prepared using spin-columns or acid guanidium-phenol- chloroform (AGPC) extraction methods contains small amount of genomic DNA. Any contaminating genomic DNA will be amplified along with cDNA, especially when primer pairs are designed within the same exon or from pseudogenes. Amplification from genomic DNA can result in qualitative and quantitative inaccuracies. The genomic DNA degradation and revers transcription steps can be achieved sequentially without purification or heat inactivation of DNase I.

SensiFAST™ cDNA Synthesis Kit - Bioline

® Cell Lysis & RT Kit for qPCR (Code No. SCQ-101) consists of "Lysis Reagents" and "RT Reagents" for synthesis of cDNA templates for real-time PCR assays. "Lysis Reagents" can be used to prepare cell lysates containing RNAs that can be used as templates for reverse transcription.
"RT Reagents" can be used to carry out reverse transcription, and are optimized for efficient cDNA synthesis from crude lysates. The synthesized cDNA can be used as a template for two step real-time PCR. The cell lysate prepared by "Lysis Reagents" can be applied to one-step real-time PCR.

cDNA Synthesis & RT-PCR | NEB

Since first-strand cDNA synthesis is less than 100% efficient, I doubt whether it can make the smear many times brighter than the mRNA in the original total RNA.)

2) Will there be a difference in result if an oligo(dT) or anchored oligo(dT) primer is used versus a random hexamer primer?

cDNA Synthesis Kits | Biocompare Editorial Article

SMART (Switching Mechanism at 5’ End of RNA Template) is a unique technology that allows the efficient incorporation of known sequences at both ends of cDNA during first strand synthesis, without adaptor ligation. The presence of these known sequences is crucial for a number of downstream applications including amplification, RACE, and library construction. While a wide variety of technologies can be employed to take advantage of these known sequences, the simplicity and efficiency of the single-step SMART process permits unparalleled sensitivity and ensures that full-length cDNA is generated and amplified.

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