34 results about "HpaII" patented technology

The invention discloses a PCR detection method for identifying germplasms of four populations of Pelodiscus sinensis, a primer group and a kit. The primer group comprises primers of SEQ No.1 / SEQ No.4, SEQ No.2 / SEQ No.5 and SEQ No.3 / SEQ No.6. The kit comprises a 2*TaqPCR premixed liquid, a 5U / mul enzyme BglI and 10*digestion buffer liquid A, a 5U / mul HpaII and 10*digestion buffer liquid B, a 5U / mul ClaI and 10*digestion buffer liquid C, 10mg / ml BSA, a contrast DNA template and a 10pM amplimer. The method comprises the following steps: carrying out PCR amplification of extracted DNA through using the primer group, digesting to obtain a digestion product, carrying out agarose gel electrophoresis, and comparing the obtained electrophoresis result with a standard electrophoretogram to obtain a result. The detection method has the advantages of simplicity, good reaction stability and repeatability, and cost saving.

The last step of synthesis of 2,3-butanediol is that acetoin reductase performs catalytic reduction on acetoin, 2,3-butanediol is synthesized, the reaction requires participation of reduced coenzyme NADH (nicotinamide adenine dinucleotide hydrogen), and thus the high-low level of intracellular coenzyme NADH is an important factor affecting conversion of acetoin to 2,3-butanediol. In order to further improve the synthesis capability of strain 2,3-butanediol and reduce accumulation of byproduct acetoin, HpaII-fdh fragments are inserted into a gene nox, a fragment nox::HpaII-fdh is obtained and guided into bacillus amyloliquefaciens B10-127, and recombinant bacillus amyloliquefaciens (renamed as N delta F) with the fdh gene inserted into the gene nox on a genome is obtained in a homologous recombination principle. Fermentation performance detection of 2,3-butanediol is performed on original bacteria and recombinant bacteria N delta F. A result proves that by comparison with the original bacteria, the yield of the recombinant bacteria 2,3-butanediol is improved by 18.9%, and the yield of the byproduct acetoin is reduced by 70.8%. Accordingly, by means of the strategy, the yield of 2,3-butanediol can be improved, and accumulation of the byproduct acetoin can be reduced.

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.