Description:Total RNA used for cDNA synthesis is isolated by modified guanidine thiocyanate techniques. 11ug total RNA was primed by an oligo dT primer and reverse transcribed by MMLV reverse transcriptase in 40ul final volume. RT Reaction stopped by heating at 65ºC for 10 minutes. The cDNA is in 1X RT buffer. (1X RT Buffer: 50mM Tris-CI, pH 8.3, 75mM KCI, 3mM MgCI2, 10mM DTT). The estimated cDNA concentration is about 2.5ng/ul. 1ul cDNA is sufficient for one PCR reaction.
Quality Control:1. The integrity of the RNA used for cDNA synthesis is examined by visual inspection for the presence of intact bands of 18s and 28s ribosomal RNA when electrophoreses on a denaturing agarose gel. The quality and purity of total RNA were tested by spectrophotometer. A260/280 is between 1.8 and 2.0 (detected in 10mM Tris-Cl, pH 7.5). The ratio of 28S/18S is ≥1
2. The RNA used for cDNA synthesis is treated by DNase I, and is tested as DNA free RNA by PCR
3. The synthesized cDNA was 5’ selected to ensure its full length. The cDNA was used as template for PCR amplification of b-actin gene and an 838bp b-actin band was visualized on 2% agarose gel. b-Actin control primer is included (T5595-0010).
Applications:Immediate PCR Amplification of known genes. Verification of genetic mutation. Comparison of a specific gene between different tissues. Analysis of mRNA alternative splicing. Gene cloning and target sequencing
Unit:1ul cDNA is sufficient for one PCR reaction.
Storage and Stability: Store cDNAs at -20°C
Unique Features of cDNA:PCR Ready First Strand cDNA is made from our high-quality total RNA. Each cDNA is a premade, tissue-specific “pool” of first strand cDNA from which full-length genes can be amplified by using sets of gene-specific primers. They can also be used to study tissue-specific gene expression and to find not only polymorphic forms of mRNA but also mRNA belonging to a multigene family. The following features have been integrated into our cDNA products to ensure their superior quality.
1. Three formats of cDNA products are availableThere are three types of cDNA products to choose from: PCR ready first strandcDNA, tumor/adjacent normal paired cDNA, and cDNA panels.
2. RNAs with high quality are used for cDNA synthesisThe cDNA synthesis will not be successful using degraded or contaminated RNA, because degraded RNA will not generate full-length cDNA, and contamination may inhibit the synthesis of cDNA. The RNAs isolated by BioChain’s proprietary technology are pure and intact from primary tissues. These RNAs therefore ensure a very high quality of cDNAs.
3. Efficient reverse transcription is guaranteedAn incorporation test is integrated into our quality control procedure for cDNA synthesis. Every cDNA product must pass a standard high incorporation.
4. cDNA is full lengthAll cDNA products are reverse transcribed with oligo dT primer, and 5’ selected. With the cDNAs as template, 5’ and 3’ regions of Clathrin, a relatively low abundance gene, are amplified by PCR. The generation of PCR products indicates the gene is not only expressed but is also full length. Since the size of clathrin is over 6kb, any genes less than 6kb are guaranteed to be full length.
5. The cDNA gene expression profile is the same as the mRNA profileThe expression patterns of any genes from different tissue cDNAs are the same as total RNA or mRNA. The cDNA can be used not only for amplification of a gene fragment but also for gene quantification.
6. Placenta cDNA is established as a gene expression standardAny gene expression data generated anywhere or anytime can be compared with this standard.
7. The largest selection of tissue cDNAs on the marketUnited States Biological provides tissue cDNAs from different human adult and fetal normal tissues, diseased and tumor tissues, mouse, rat, monkey, and plant tissues.
8. Tumor/adjacent normal tissue cDNA pairs and cDNA panels are availableTumor/adjacent normal cDNA pairs and cDNA panels combine high quality cDNAs selected from almost 200 different human adult and fetal normal tissues, diseased and tumor tissues, mouse, rat, monkey, and plant tissues. Each panel contains a variety of cDNAs from different tissues. All cDNAs in a panel are made under optimized and consistent conditions, ensuring that cDNA gene expression profiles are the same as those of total RNA.
Our cDNA panel is provided ready for PCR quantification of gene expression. Our cDNA panels are perfect for analysis of poorly expressed genes that are not detectable by Northern analysis or RNase protection assay. The cDNA panels are also ideal for identification of gene expression in some rare tissues.
仕様
Size：40Tests
Source Antigen：Human
Grade：Molecular Biology Grade
Form：Supplied as a liquid in 1X RT Buffer: 50mM Tris-Cl, pH 8.3, 75mM KCl, 3mM MgCI2, 10mM DTT.
