A method for expressing restriction endonucleases efficiently through the protection of methylases

Abstract

The invention discloses a method for protecting efficient recombinant expression of restriction enzymes by virtue of methylase. The method is characterized in that the methylase is M.Sss I; and the restriction enzymes D are selected from AatII, AciI, AclI, AfeI, AgeI, AscI, AsiSI, AvaI, BceAI, BmgBI, BsaAI, BsaHI, BsiEI, BsiWI, BsmBI, BspDI, BspEI, BsrFI, BssHII, BstBI, BstUI, BtgZI, ClaI, EagI, FauI, FseI, FspI, HaeII, HgaI, HhaI, HinP1I, HpaII, Hpy99I, HpyCH4IV, KasI, MluI, NaeI, NarI, NgoMIV, NotI, NruI, Nt.BsmAI, Nt.CviPII, PaeR7I, PluTI, PmlI, PvuI, RsrII, SacII, SalI, SfoI, SgrAI, SmaI, SnaBI, TliI, TspMI, XhoI, XmaI and ZraI. According to the method provided by the invention, the broad-spectrum protective methylase M.Sss I is directly adopted in a recombinant engineering bacterium construction process, without conducting complex methylase gene screening on a single restriction enzyme, so that the method is broad in application scope and is capable of greatly simplifying a recombinant expression process.

Description

technical field [0001] The invention belongs to the field of bioengineering, in particular to a method for efficiently recombining and expressing a restriction endonuclease using the protection of methylase. Background technique [0002] Restriction endonucleases are indispensable and important tools for contemporary genetic engineering research. They are derived from different microorganisms and are weapons for bacteria to defend against foreign virus invasion. In the 1960s, scientists speculated that there is a restriction-modification system (R-M system) in bacteria, which includes two types of enzymes, namely restriction endonucleases and DNA methylases, and the former is responsible for degradation into prokaryotic cells. The exogenous DNA of the cell, which methylates the cell's own DNA, thereby protecting the cell's DNA from being degraded by restriction endonucleases in the cell. [0003] The method for producing restriction endonucleases in the prior art is mainly ...

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Problems solved by technology

[0003] The method for producing restriction endonucleases in the prior art is mainly to clone restriction-modified genes into plasmids, to achieve high expression of target proteins by the high copy number of plasmids, and to use specific formazan corresponding to restriction endonucleases. However, the purification procedure of this method is cumbersome, the yield of restriction endonuclease is low, and the production cost is high. The obtained methylation-protected strain can only specifically protect the host DNA from a certain restriction endonuclease. Enzymatic cleavage, narrow range of applications

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