The restriction endonuclease hydrolyzes the phosphodiester bond in the nucleic acid strand in an endogenous manner, and the resulting DNA fragment has a P' at the 5' end and an OH at the 3' end.
1. Types of restriction enzymes According to the recognition and cleavage properties of restriction enzymes, the catalytic conditions and whether they have modified enzyme activities can be divided into three categories: I, II and III.
The first type (type I) restriction endonuclease recognizes a specific nucleotide sequence that arbitrarily cleaves the DNA strand at a position far from the recognition site, but the nucleotide sequence of the cleavage is not specific and random. of. Such restriction enzymes are of little use in DNA recombination techniques or genetic engineering and cannot be used to analyze DNA structures or clone genes. Such enzymes are, for example, EcoB, EcoK and the like.
The third type (type III) restriction endonuclease also has a specific recognition sequence that cleaves the double strand at a fixed position of several nucleotide pairs next to the recognition sequence. But these few nucleotide pairs are not specific. Therefore, a certain length of DNA fragment produced by this restriction endonuclease cleavage has various single-stranded ends. Therefore, it cannot be applied to gene cloning.
The third type (type II) restriction endonuclease is commonly referred to as DNA restriction endonuclease.
They recognize the specific sequence of double-stranded DNA and cleave within this sequence to produce specific DNA fragments;
The type II enzyme has a small molecular weight and only requires Mg2+ as a cofactor for the catalytic reaction, and the recognition sequence is generally an inverted repeat sequence of 4 to 6 base pairs;
Type II endonuclease cleaves double-stranded DNA to produce three different nicks - 5' end overhangs; 3' end overhangs and ends.
Thanks to the discovery and application of restrictive endonucleases, people can purposely transform genetic DNA in vitro, which greatly promotes the prosperity and development of molecular biology.
Sticky ends: are staggered cuts, resulting in the formation of two single-stranded ends, the nucleotide sequences of which are complementary and form hydrogen bonds, so called sticky ends.
For example, the recognition order of EcoRI is:
5'... G|AATTC ......3'
3'... CTTAA|G ...... 5'
Generated after cutting at the position of the staggered cut on the double chain
5'...G AATTC...3'
3'...CTTAA G...5'
Each has a single-stranded end, the two single strands being complementary, and the cleavable phosphodiester bond and hydrogen bond can be "bound" by the action of DNA ligase.
Flat end:
Another type of type II enzymatic cleavage is the cleavage of the duplex at the same position, resulting in a blunt end. For example, the recognition location of EcoRV is:
5'... GAT|ATC ...... 3'
3'... CTA|TAG ...... 5'
Formed after cutting
5'... GAT ATC ...... 3'
3'... CTA TAG ...... 5'
This end can also be linked by DNA ligase.
2. The nomenclature of restriction endonucleases indicates the species name of the host bacteria using the first letter of the generic name and the first two letters of the species name. For example, E. coli is represented by Eco, so it is in italics.
One letter is used to represent the strain or type, such as Hemophilus influenzae Rd strain, d, ie Hind.
If a particular host strain has several different restriction and modification enzymes, it is represented by Roman numerals, such as HindI, HindII, HindIII, and the like.
Non-Inactivated Disposable Virus Sampling Tube
This disposable Non-Inactivated virus sampling tube is an upgrade from the Hank's solution. Additional various components, such as BSA, HEPES, amino acids, cryoprotectant, etc., are added to enhance the virus integrity. It can be used for nucleic acid extraction of virus, mvcoplasma chlamydia and ureaplasma samples and later virus isolation.
This disposable non-inactivated virus sampling tube consists of fibrous swab and culture medium.The medium contains antibiotics and antimycotics for the purpose of inhabitation of bacteria and veasts overgrowth. It helps to maintain the cellular integrity and preserve the viruses, mycoplasma, chlamydiae and ureaplasma.
When collect specimens with this disposable Non-inactivated virus sampling tube, tube can be transported to the laboratory at 2-8ºC within 2 days. If it can not be delivered to the laboratory within 48hours, it should be stored at-70ºC or below, and ensure that the collected specimens are delivered to the laboratory within 1 week. Avoid repeated freeze-thaw cycles. For best performance, complete testing within 24 hours.
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