What is subtractive DNA hybridization?

Subtractive hybridization is a technique used to isolate a DNA segment that is missing from one particular sample of DNA. Obviously, a second DNA sample that contains the fragment of interest is necessary. Suppose that a hereditary defect is due to the deletion of the DNA for a particular gene.

How is subtractive hybridization useful?

The advantages of subtractive hybridization include the need, particularly when combined with poly(A) RT-PCR, for only very small amounts of mRNA, the ability to detect rare mRNAs, representing as little as approximately 0.01% of the total number of mRNA species present, and the ability to clone novel genes.

What does suppressive subtractive hybridization do?

Suppression subtractive hybridization (SSH) is a widely used method for separating DNA molecules that distinguish two closely related DNA samples. Two of the main SSH applications are cDNA subtraction and genomic DNA subtraction.

What is subtractive cDNA library?

Subtractive cDNA libraries are produced using a proprietary technique which relies on the removal of dsDNA formed by hybridization between a control and test sample, thus eliminating cDNAs of similar abundance and retaining the transcripts which are differentially expressed or variable in sequence.

What is cDNA in biology?

Complementary DNA (cDNA) is a DNA copy of a messenger RNA (mRNA) molecule produced by reverse transcriptase, a DNA polymerase that can use either DNA or RNA as a template. From: Encyclopedia of Genetics, 2001.

What is a subtracted library?

A subtracted cDNA library contains cDNA clones corresponding to mRNAs present in one cell or tissue type and not present in a second type.

What is subtraction library?

Subtraction-hybridization cDNA libraries (1, 2, 3, 4) are libraries enriched for sequences representing mRNAs whose expression in one biological source (e.g., tissues, cell lines) is different than in a second source.

What is full length cDNA?

Definition. cDNA is a ‘complementary’ or ‘copy‐DNA’ that is generated in vitro from cellular mRNA (messenger RNA). Full‐length cDNAs contain the complete sequence information of their respective mRNA templates.

What is cDNA cloning?

cDNA cloning is isolating and amplifying a single, self-replicating organism that includes within its DNA, a cDNA that is of interest to the experimenter.

How stable is cDNA?

The cDNA (prior to cDNA amplification) can be safely stored overnight at 4˚C or -20˚C. If the sample must be stored for a longer time, add 0.5 µl of 10X NEBNext Cell Lysis Buffer to the cDNA (after RT instead of during the PCR step).

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