80 results about "Chemo enzymatic" patented technology

The invention relates to heparinase III with high activity on 6-sulfated heparin and application of the heparinase III. The amino acid sequence of the heparinase III (BeHepIII) is as shown in SEQ ID NO. 2, and the nucleotide sequence of the coding gene is as shown in SEQ ID NO. 1. The invention discovers and discloses a novel heparinase III (BeHepIII) for the first time, the heparinase III is derived from Bacteroides eggerthii, has a higher recombinant expression level in escherichia coli, has very high degradation activity on heparin containing a 6-site sulfated structure, and particularly has the enzyme activity which is 93.5 times of that of commercialized heparinase FhHepIII at present, and the heparinase III can be used for degrading heparin containing 6-site sulfated structure. Therefore, the heparinase III (BeHepIII) provided by the invention has a wide application potential in heparin degradation. In addition, by using heparinase III (BeHepIII), directional excision of a reducing end of heparin oligosaccharide synthesized by a chemical enzyme method is realized, and complete chemical enzyme method synthesis of a pentasaccharide structure with anticoagulant factor Xa activity is successfully realized.

The invention discloses a chemical-enzymatic coupling synthesis method of oseltamivir phosphate, which comprises the following steps: by taking monasic acid OS-01 as a substrate, sequentially carrying out S1 esterification, S2 ketalation, S3 selective ring opening, S4 TEMPO oxidation, S5 primary transaminase reaction, S6 acetylation, S7 secondary transaminase reaction and S8 phosphorylation reaction to obtain the final product oseltamivir phosphate OS-09. According to the method, the step of sodium azide required by a traditional route is avoided, the production safety and the industrialization possibility are enhanced, two different transaminases are used for respectively constructing different chiral centers, the steps of protecting groups and deprotecting groups on the traditional route are not needed, the overall route is short, the used chemical reagents are few, the yield is high, and the method is suitable for industrial production. The industrial requirements of environmental protection and low cost are met.

This invention relates to the fields of agricultural product processing and bio-feed technology, specifically to a method and application for detoxifying corn steep liquor. The method includes: subjecting a mixture of corn steep liquor, a composite color-protecting agent, and a composite catalyst to a first reaction; allowing the reaction system after the first reaction to stand in a mixture with a composite adsorbent; subjecting the mixture containing the reaction system after the stand-in reaction and a composite enzyme preparation to stand-in treatment, mixing treatment, and enzymatic hydrolysis; and subjecting the mixture containing the reaction system after the enzymatic hydrolysis and an activated composite microbial agent to anaerobic fermentation. This method employs a multi-stage chemical-enzyme-fermentation linkage process, achieving efficient detoxification of corn steep liquor while significantly improving flavor and nutrition. This method synergistically removes mycotoxins and sulfites, and enhances palatability and probiotic value through fermentation transformation of off-flavor substances and enrichment of small peptides and lactic acid. The process is simple and requires no adsorbent separation.

The invention relates to the technical field of preparation of electrochemical sensors, and particularly discloses a preparation method of a novel electrochemical enzyme sensor for profenofos pesticide detection based on an enzyme inhibition action principle, and the preparation method comprises the following steps: S1, treating a glassy carbon electrode to obtain a bare glassy carbon electrode GCE; s2, obtaining a composite material modified electrode Au50Ag2 (at) G / GCE; s3, an enzyme-chitosan composite modified electrode KbE-CS / Au50Ag2 (at) G / GCE is obtained; and S4, finally obtaining the profenofos electrochemical enzyme sensor NF / KbE-CS / Au50Ag2 (at) G / GCE based on the enzyme inhibition effect. According to the present invention, the white kidney bean esterase derived from plants is adopted as the detection enzyme source, and the electro-catalysis amplification effect of the Au50Ag2 (at) G composite material is adopted to construct the novel electrochemical enzyme sensor, and the method is the profenofos pesticide detection technology based on the enzyme inhibition effect principle;

The invention belongs to the technical field of biological medicine and disease prevention and treatment, and particularly relates to an SSEA4 conjugated virus-like particle anti-cancer vaccine as well as a preparation method and application thereof. Specifically, according to the vaccine, an SSEA4 antigen is efficiently synthesized through a chemical enzyme method, then the SSEA4 antigen is covalently coupled with a virus-like particle (VLP), the SSEA4 antigen and the VLP are combined with a monophosphoryl lipid A adjuvant with high safety, an organism can be induced to generate strong and lasting SSEA4-resistant humoral immune response, the generated antibody can specifically recognize and kill SSEA4 positive tumor cells, and the SSEA4 positive tumor cells can be effectively killed. Meanwhile, good safety is shown in an animal model, a new solution is provided for immunotherapy of SSEA4 positive tumors, and therefore the SSEA4 positive tumor immunotherapy polypeptide has wide application prospects and market value.

The invention belongs to the field of feed protein processing, and particularly relates to a method for improving the bioavailability of animal feed protein through chemical enzymolysis. The method comprises the following steps: mixing a compound protein raw material with water to form a suspension; sodium tripolyphosphate is added for phosphorylation pretreatment, so that protein is subjected to specific modification; after the pH is adjusted to be acidic, pepsase and acid phosphatase are added at the same time for an enzymolysis reaction; and after the reaction is finished, performing enzyme deactivation and drying to obtain the modified feed protein. According to the method, a specific enzyme cutting site is created on a protein molecule through chemical phosphorylation, then directional hydrolysis is achieved through efficient recognition of pepsin on the site, and meanwhile dynamic regulation and control of the hydrolysis process are achieved by means of the dephosphorylation effect of acid phosphatase. The obtained product has the outstanding advantages of high content of small peptides, good digestive absorbability, low bitterness, strong biological activity and the like, and the bioavailability of feed protein is remarkably improved.

The invention relates to a chemoenzymatic synthesis method of a high-glucose-type homogeneity mucoprotein core structural domain. The method comprises the following steps: firstly, catalytically synthesizing a key alpha-linked glycosylated amino acid block Fmoc-GalNacalpha-Ser / Thr-OH at a low temperature (-20 DEG C to 0 DEG C); then assembling the building blocks on a repeated polypeptide skeleton rich in proline, serine and threonine at accurate intervals (3-5 amino acid residues) through a solid-phase peptide synthesis technology to form a'sugar chain growth scaffold '; then, a series of high-specificity glycosyl transferases are used for carrying out sequential and directional sugar chain enzymatic extension on the scaffold, and a uniform core structure (such as a core type 3) is constructed; and finally, performing high-efficiency purification by a three-step series method of size exclusion chromatography, ion exchange chromatography and lectin affinity chromatography. The technical bottleneck of inhomogeneity (lt, 70%) of glycoforms in a traditional method is overcome, the mucoprotein core fragment with glycoform homogeneity larger than or equal to 95% and bioactivity highly similar to that of natural mucoprotein can be prepared on a large scale, and the method has wide application prospects in the biomedical fields of mucous membrane protective agents, drug delivery carriers and the like.

This invention discloses a class of pentacyclic triterpenoid derivatives with diverse B-ring modifications, their preparation methods, and applications. This invention utilizes a chemical enzymatic method to expand the chemical space of the B-ring in pentacyclic triterpenoids, obtaining a series of pentacyclic triterpenoid derivatives with diverse B-ring modifications. Pharmacological activity screening of these pentacyclic triterpenoid derivatives revealed their good tumor-suppressive activity, providing a promising application prospect for the prevention and treatment of tumors. This invention achieves the efficient synthesis of pentacyclic triterpenoid derivatives with diverse B-ring modifications, structures that are difficult to obtain using traditional chemical methods. Systematic antiproliferative activity screening identified candidate molecules with broad-spectrum and highly efficient inhibitory effects against various human tumor cell lines. These molecules show good drug development potential, providing novel lead compounds for the innovative development of anticancer drugs.

The application uses cheap and easily available hexose acid as a starting material, first obtains two precursors of 2,5-furan dicarboxylic acid synthesis, namely hexahydroxy acid and 4-deoxy-5-dehydro-hexahydroxy acid through enzyme catalytic reaction, and then 4-deoxy-5-dehydro-hexahydroxy acid is dehydrated under acidic conditions to obtain 2,5-furan dicarboxylic acid.

The application discloses a cytochrome P450BM3 mutant, a carrier, microorganisms and application thereof, and belongs to the technical field of chemical enzyme synthesis.The application discloses a synthetic route of a key drug intermediate dehydro-nandrolone acetate synthesized by a chemical enzyme method, wherein enzyme catalysis and chemical catalysis are included.The enzyme catalysis includes a cytochrome P450 mutant and 17beta-steroid reductase, and a one-pot two-step method is used to catalyze acid to be removed to generate 7beta-hydroxynandrolone.Combined with chemical catalysis, the 7beta-hydroxynandrolone is subjected to dehydration esterification to generate the dehydro-nandrolone acetate.The synthetic route has short steps, high yield and the ability to reduce the production cost of related steroid drugs.

Presented are kainoids, kainoid precursors, and kainoid derivatives and synthetic schemes for producing kainoids, kainoid precursors, and kainoid derivatives. The synthetic schemes for producing kainoids can make use of kainoid synthases, for example, in cyclizing a kainoid precursor to produce a kainoid. The kainoids can include an alkyne group 5 to facilitate the production of kainoid derivatives.

Disclosed is a method for forming selenoneine or analogs thereof. The method may include phosphorylating sodium selenide to a selenophosphate, using adenosine triphosphate (ATP) and at least a first protein, generating a selenosugar by converting the selenophosphate using at least a second protein in the presence of a common sugar donor, and forming selenoneine or an analog thereof by combining the selenosugar with N,N,N-trimethyl-L- histidine or analogs thereof using at least a third protein. The method may include combining SenA, SenB, and SenC in an aqueous buffer at neutral pH and ambient temperature, and allowing SenA, SenB, and SenC to form selenoneine or an analog thereof in the presence of ATP, a common sugar donor, and sodium selenide.

The present invention discloses a method for mechanochemical enzyme-coupled lignin degradation. The specific operation process is to place lignin powder, a solid-phase reagent, and zirconium oxide beads in a polytetrafluoroethylene ball mill, perform a co-grinding reaction in the ball mill, add an enzyme after ball milling, add a solvent for stirring, and reflux reaction. After the reaction, the mixture is dissolved in water and extracted with ethyl acetate. The insoluble residue is centrifuged, washed with ethyl acetate, and vacuum dried for detection and analysis. The present invention has the advantages of a short process route, low reagent usage, simple operation, low cost, low pollution, low molecular weight of the obtained product, and good degradation efficiency. It is a lignin degradation method with good promotion and application prospects.

The invention belongs to the crossing field of a biosensing technology and efficient screening of natural medicines, and particularly relates to an electrochemical enzyme biosensor based on synergistic interaction of a nano material and application of the electrochemical enzyme biosensor in rapid screening of anti-gout active components. In order to solve the problems of low efficiency, high cost, dependence on in-vitro biochemical detection, high false positive rate and the like in the traditional drug screening technology, high-throughput and high-sensitivity detection of anti-gout active components in natural products is realized by innovatively constructing a xanthine oxidase (XO) immobilized nano-composite sensing interface.

The present invention provides a chemoenzymatic synthesis method for blood group trisaccharide, which includes: dissolving a substrate, GDP-Fuc, and magnesium chloride in water, adjusting the pH of the solution, adding the enzyme α1,2-FucT, and stirring for reaction to obtain a first product reaction solution; adding a de-enzyme solvent, standing, centrifuging and filtering, taking the supernatant, and evaporating to remove the de-enzyme solvent to obtain an aqueous solution of the first product; adding a glycosylated product, adjusting the pH after dissolution, and adding a glycosyltransferase to obtain a second product reaction solution; using a de-enzyme solvent to remove the glycosyltransferase, standing, centrifuging and filtering, taking the supernatant, and evaporating to remove the de-enzyme solvent to obtain an aqueous solution of the second product; separating and purifying the aqueous solution of the second product, and freeze-drying to obtain a solid product; dissolving the solid product, adding Pd / C, introducing hydrogen gas, stirring and filtering to remove Pd / C, concentrating and freeze-drying to obtain the blood group trisaccharide. This method provides a new technical idea for the chemoenzymatic synthesis of blood group trisaccharide, which is conducive to popularization and application and solves the problems of low yield and high price.

The invention firstly discloses a method for synthesizing the spraadine A and the 3-OH spraadine A by utilizing a chemical-enzyme method. The synthetic route mainly comprises three steps: 1) synthesizing (-)-beta-CPA through a novel chemical enzyme method synthetic route; (2) catalyzing the (-)-beta-CPA in vitro by using oxidation cycloaddition enzyme CpaO to carry out intramolecular asymmetric oxidation [3 + 2] cycloaddition reaction to generate (-)-alpha-CPA; and (3) by taking the (-)-alpha-CPA as a substrate, synthesizing alpha-CPA natural products in a high oxidation state, such as speradine A, 3-OH speradine A and the like, through chemical modification. The invention has important theoretical and practical significance for three-dimensionally and specifically preparing or synthesizing a series of high-oxidation-state alpha-CPA natural products, and provides a leading compound for the development of anti-cancer drugs.

Provided herein are synthetic methods and intermediates useful in the preparation of polyketides, including 12-membered ring macrolides. These synthetic methods and intermediates provide scalable access to polyketides, including polyketides with potent bioactive splicing modulator activity and therapeutic potential, including treating various types of cancer. The polyketides include 12-membered ring macrolides, including Compound A, Compound B, and pladienolides, including pladienolide A, pladienolide B, pladienolide C, pladienolide D, pladienolide E, pladienolide F, and pladienolide G.

The invention discloses a method for extracting ganoderma lucidum polysaccharide by a microwave-assisted green chemical-enzyme method and application, which comprises the following steps: taking ganoderma lucidum sporocarp powder, adding cellulase and water, uniformly mixing, then adding 30% hydrogen peroxide, uniformly mixing, adjusting the pH value to 3-7, and carrying out microwave-assisted extraction for 40-120 minutes under the conditions that the power is 400-800W and the temperature is 50-90 DEG C; and centrifuging, taking supernate, concentrating to 30-50% of the original volume, and freeze-drying to obtain the ganoderma lucidum polysaccharide. The method is high in ganoderma lucidum polysaccharide extraction efficiency and more complete in ganoderma lucidum polysaccharide activity retention. The ganoderma lucidum sporocarp polysaccharide product is free of peculiar smell and good in sensory property, the SOD activity in C.elegans under the hydrogen peroxide stress state can be improved by 33.76%, the CAT activity can be improved by 229.36%, and the MDA content can be reduced by 85.65%.

A field of glycoproteomics for identifying novel cell surface O-linked glycopeptide epitopes. The disclosure relates to methods for identifying O-linked glycopeptides, more specifically, Tn- or SiaTn-antigen O-glycosylation sites on cell surface proteins, the method comprising a combined workflow of enzymatic treatments of cells or tissue samples to allow specific enrichment and labelling of Tn antigens, followed by mapping of the O-glycopeptides through LC-MS / MS. More specifically, the disclosure relates to a chemo-enzymatic method to produce labelled O-glycan-peptides from a plasmamembrane-protein extract sample, wherein Sialyltransferase (ST3Gal1 / ST3Gal3) treatment is used to protect free galactose against a follow-on treatment with Galactose oxidase, wherein the oxidized glycans are simultaneously labelled. The disclosure further relates to a tandem mass spectrometry method using an enriched TMT-labelled (Sia)Tn-antigen glycopeptides in a dual fragmentation triggered approach to resolve the identity, localization and quantification of the O-linked glycan peptides.

The present invention discloses a method for synthesizing danshensu using a chemoenzymatic cascade system. The first enzymatic cascade system or the second enzymatic cascade system is subjected to a catalytic reaction to generate (R)-2-hydroxy-3-(4-hydroxy-3-methoxyphenyl) propionic acid, and then a demethylation reaction is carried out using hydrobromic acid to finally generate D-danshensu. The technical solution provided by the present invention synthesizes danshensu using vanillin as a raw material. Vanillin has a low cost and stable chemical properties; the reaction conversion rate of the chemoenzymatic cascade system for synthesizing danshensu is high, the conversion rate of vanillin can reach 100%, and it has excellent chemoselectivity. The yield of danshensu reaches 90%, all of which are D-danshensu, and no other by-products are generated, showing good application prospects.

The invention discloses an olefin reductase SeER mutant and application thereof in synthesis of chiral drugs, and provides a series of novel olefin reductase SeER mutants capable of efficiently and asymmetrically reducing beta-alkyl / aryl-beta-cyanoacrylate compounds, and the olefin reductase SeER mutants have high conversion rate (gt; 99% conversion rate), good enantioselectivity (gt; 99% ee), the substrate range is wide (gt; according to the present invention, the screened olefin reductase SeER mutant can be successfully synthesized into the chiral GABA derivative drugs such as pregabalin and brivaracetam through the chemical-enzyme coupling method, such that the overall reaction yield is improved, and the route is simple;

Disclosed are intermediate compounds (i.e., oligonucleotide fragments), or pharmaceutically acceptable salts thereof, useful in the preparation of RNAi agents. In addition, the present invention discloses methods of making single stranded oligonucleotides by linking two or more intermediate compounds described herein by a hybrid chemical-enzymatic pathway.

A uridine diphosphate-N-difluoroacetylglucosamine, a preparation method therefor and a use thereof. The uridine diphosphate-N-difluoroacetylglucosamine has a similar chemical structure to the natural substrate UDP-GlcNAc, the difference being that the 2-position N-acetyl group thereof is replaced by an N-difluoroacetyl group. The preparation method is as follows: (1) preparing difluoroacetylglucosamine; (2) using difluoroacetylglucosamine as a substrate to prepare uridine diphosphate-N-difluoroacetylglucosamine. A novel, highly active, non-hydrolyzable and artificially synthesizable artificial donor sugar uridine diphosphate-N-difluoroacetylglucosamine (UDP-GlcNDFA) is provided. Compared with the existing donor sugar UDP-GlcNTFA, UDP-GlcNDFA has higher catalytic activity for glycosyltransferases, is stable and not easily hydrolyzed, and can be used as a donor substrate for chemoenzymatic synthesis of heparin oligosaccharide scaffolds, heparin oligosaccharide intermediates and heparin oligosaccharides, as well as hyaluronic acid oligosaccharides, glycoconjugates (glycosylated polypeptides) and polysaccharide antigens.

The invention discloses a method for synthesizing ergothioneine based on a chemical-enzyme method, and belongs to the technical field of biology. The method comprises the following steps: firstly, synthesizing histidine betaine by using a chemical method, and carrying out selective trimethylation reaction in an alkaline aqueous solution by using methyl iodide as a methyl donor, so that the final concentration of histidine betaine is 77g / L, and the conversion rate is 93%; then, histidine betaine sulphide enzyme EanB from an anaerobic biological argillobiosis source is selected, a histidine betaine sulphide enzyme mutant F97A / T387M / E277K / N354Q / F441S / S335K is constructed through truncation transformation and mutation, and histidine betaine is directly vulcanized to synthesize ergothioneine. According to the present invention, through the cascade reaction of the chemical-enzyme method, the synthesis from the histidine to the ergothioneine is successfully achieved, the final ergothioneine yield is 33.5 g / L, and the conversion rate is 95%;

The application belongs to the technical field of biology and specifically relates to a compound for treating myocardial infarction. The chondroitin sulfate oligosaccharide in the application is extracted from natural chondroitin sulfate, and after being treated into an oligosaccharide chain through a chemical enzyme method, experiments prove that the chondroitin sulfate oligosaccharide is helpful to the survival of myocardial cells. Compared with other similar functional products, the glycopeptide polymer hydrogel formed after the introduction of a polymer skeleton overcomes the problem of batch difference of natural products, has a clear structure, and the hydrogel itself has excellent biological activity, and does not need to encapsulate other active ingredients such as drugs. The effect of supporting the infarct area of the ventricular wall and promoting the survival of myocardial cells is obvious, the myocardial infarction area can be effectively limited, the thickness of the infarct area of the ventricular wall is increased, the damage of the heart infarction to the heart is reduced, and the application has a good application prospect.

The application discloses an olefin reductase, a coding gene, a carrier, an engineering bacterium and application thereof, wherein the olefin reductase SeER mutant is obtained by single-point or multi-point site-directed mutation or site-directed saturation mutation of the 38th, 117th, 248th, 293rd and 373rd sites of the amino acid sequence shown in SEQ ID NO. 2. The application provides a series of new olefin reductase SeER mutants capable of efficiently stereocomplementary reduction of olefin compounds to prepare chiral compounds, and successfully synthesizes chiral GABA derivative drugs such as phenibut, baclofen, tolperisone and pregabalin through a chemical-enzyme coupling method. Compared with existing methods, the strain can flexibly regulate the configuration of the product, and has high stereoselectivity and conversion rate.

The present invention provides a method for preparing dehydroepiandrosterone by a chemical-enzymatic method, belonging to the technical field of pharmaceutical compounds. In the present invention, water, glacial acetic acid and sodium D-ascorbate are mixed and then tert-butanol, a strong base catalyst and 4-androstenedione are sequentially added for reaction to obtain 5-androstenedione; a phosphate buffer system is obtained by mixing tert-butanol with a phosphate buffer solution; 5-androstenedione, magnesium chloride, ketoreductase, coenzyme and a coenzyme regeneration system are added to the phosphate buffer system for an enzymatic reaction, and dehydroepiandrosterone is obtained. The target product is synthesized in two steps by the combined chemical-enzymatic method in the present invention, and the yield of the product is as high as 95%, and the purity is above 99%. The synthesis method of the present invention effectively solves the problems of complex chemical process and difficult post-treatment of biological method, not only simplifies the process, but also improves the yield and purity of DHEA, and can realize large-scale production.

The invention relates to uridine diphosphate-N-difluoroacetyl galactosamine as well as a preparation method and application thereof. The chemical structure of the UDP-GalNDFA is similar to that of a natural substrate UDP-GalNAc, and the core difference is that N-acetyl at the site 2 of the UDP-GalNDFA is replaced by N-difluoroacetyl. The preparation method comprises the following steps: (1) preparing difluoroacetyl galactosamine; and (2) preparing the UDP-GalNDFA by taking the difluoroacetyl galactosamine as a substrate. As a novel artificial donor sugar, the UDP-GalNDFA has the characteristics of high activity, difficulty in hydrolysis and capability of being artificially synthesized. Compared with the existing donor sugar UDP-GalNTFA, the glucose UDP-GalNTFA has higher catalytic activity on glycosyl transferase, has stronger stability, is not easy to hydrolyze, and can be used as a donor substrate for synthesizing a chondroitin oligosaccharide skeleton, a chondroitin oligosaccharide intermediate and a chondroitin oligosaccharide derivative by a chemical enzyme method.

The present application relates to a kind of high activity, high thermal stability heparin N-sulfate group transferase mutant and its coding gene and application.The amino acid sequence of the N-sulfate group transferase mutant M8 is as shown in SEQ ID NO.2, and the nucleotide sequence of coding gene is as shown in SEQ ID NO.1.The N-sulfate group transferase mutant M8 is 24 amino acids in wild-type N-sulfate group transferase Mutations.Determination of stability, compared with wild-type N-sulfate group transferase at 37 DEG C only incubate less than 1 day, all activity is lost, the N-sulfate group transferase mutant M8 of the present application has high activity and extremely strong high thermal stability, can be incubated at 37 DEG C 7 days still keep about 50% activity.The present application improves the efficiency of heparin chemical enzyme synthesis, expands the application range of N-sulfate group transferase mutant, makes it possible to synthesize heparin at high temperature, greatly promotes the application development of heparin biomimetic synthesis.

Described herein are various methods of removing uracils due to the deamination of unmethylated cytosines in an assay using engineered cytosine deaminases to deaminate methylated cytosines. The Chemoenzymatic Uracil Replacement of Nucleobases (ChURN) method includes providing a sample comprising single stranded DNA library fragments in which a cytosine deaminase has deaminated methylated cytosines; contacting the sample with an uracil DNA glycosylase (UDG) to deglycosylate uracil residues to form abasic sites having a hemiacetal formation within the single stranded DNA library fragments; contacting the sample with a reactive cytosine nucleobase analog to install a cytosine at abasic sites thru a noncanonical linkage; and subjecting the sample to polymerase chain reaction (PCR) amplification resulting in double stranded DNA corrected library fragments. The Uracil Enzymatic Removal and Substitution at Errors (U-ERASE) method includes providing a sample comprising single stranded DNA library fragments in which a cytosine deaminase has deaminated methylated cytosines, synthesizing double stranded DNA library fragments in which the second strand is tagged to facilitate its downstream degradation, treating the double stranded DNA library fragments with an uracil DNA glycosylase and an endonuclease resulting in the removal of uracil bases and single nucleotide gaps at those sites, and repairing the single nucleotide gaps through treatment with a polymerase, dCTP, and a ligase, resulting in replacement of false positive uracil bases with a mismatched base, such as cytosine. Subsequently, the second strand is selectively degraded, allowing for selective amplification of the original DNA strand via PCR.