34 results about "HpaII" patented technology

The invention relates to a genetic marker for screening the combined gene model of the fourth and tenth exon variation sites of a common disease resistance BPI (bactericidal / permeability increasing protein) gene of pigs, and belongs to the technical fields of genetic breeding and molecular marker assisted selection of pigs. The method comprises the following steps of: extracting DNA (deoxyribonucleic acid) of a pig genome of a sample to be tested, and performing PCR (polymerase chain reaction) amplification on the pig genome; analyzing the PCR product of the exon 4 through SSCP (single strand conformation polymorphism), and performing enzyme digestion on the PCR product of the exon 10 through restriction enzyme HpaII, wherein when the gene model of the pig body to be tested is a CDAB model, the pig body is strong in common disease resistance. According to the application, a molecular genetic marker with higher efficiency and accuracy is provided to molecular marker assisted selection of the pig disease resistance breeding work, and an effective molecular marker breeding means is provided for improving the common disease resistance and immune response capability of piglets. The detection method is simple in operation, low in cost and high in accuracy, can realize automatic direct detection, and takes a magnificent effect in pig breeding.

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.

A method for detecting the methylation of promoters using HpaII, a methylation-sensitive restriction enzyme. In such method, DNAs, derived from clinical samples or subjects to be diagnosed, are cut with HpaII, the cut DNAs are amplified by PCR with primers capable of amplifying CpG islands, and the presence or absence of the PCR amplification products is determined using a DNA chip for methylation detection. Unlike prior approaches, the inventive method allows the methylation of gene promoters to be detected in a simple and economical manner, and thus is useful for the diagnosis of diseases such as cancer that are characterized by methylation of gene promoters.

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 belongs to the technology field of preparation and application of pig genetic markers, and in particular relates to a genetic marker by taking pig miR-27a precursor flanking genetic fragments as a trait of the litter size of a pig, a preparation method and application thereof. According to the invention, the genetic marker which is related to the character of the litter size of the pig is obtained, wherein the genetic marker comprises the pig miR-27a precursor flanking genetic fragments and is nucleotide sequences expressed by the following sequence lists: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, and SEQ ID NO:5; a base mutation of C461-T461 is generated at 461bp of the following sequence lists: the SEQ ID NO:1, the SEQ ID NO:2, the SEQ ID NO:3, the SEQ ID NO:4, and the SEQ ID NO:5; and the mutation leads to polymorphism of PCR-HpaII-RFLP. In the design, a primer is additionally provided in the miR-27a precursor flanking genetic fragments, wherein the nucleotide sequence of the primer is expressed as SEQ ID NO:6 and SEQ ID NO:7. The invention also discloses the method for preparing the genetic marker and the application of the genetic marker in the pig marker assistant selection.

The invention relates to a method for detecting DAN methyltransferase activity based on strand displacement amplification and DNAzyme amplification. A three-function double-stranded DNA probe is designed. Methylation happening to the three-function double-stranded DNA probe is specifically recognized through DAN methyltransferase. Remaining non-methylated double-stranded DNA is specifically cut through HpaII restriction enzymes, methylated double-stranded DNA triggers a strand displacement reaction, a large amount of 8-17 DNAzyme is released, the 8-17 DNAzyme catalyzes cutting of a large number of hairpin-line molecular beacon substrates, and remarkable fluorescent enhancement is triggered. The method can sensitively detect the DAN methyltransferase activity, and the detection limit is 0.0082 U / mL. The method has potential application in research of influences of anti-cancer drugs on DAN methyltransferase activity inhibition and screening of DAN methyltransferase inhibitors.

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.