fi cation of pcr products for sequencing

... Analysis, sequencing and in vitro expression of PCR products 91 within a background of nonspecific products it will be enriched for the specific template allowing efficient ampli cation by the nested PCR ... conditions offer more definitive veri cation of product identity Some combination of approaches may be required in difficult cases Of course the most definitive confirmation of identity of a PCR product ... simplify sequencing of PCR products Primers for direct sequencing The choice of primer for direct sequence analysis depends upon how much information is available before the PCR One of the original PCR...

Ngày tải lên: 25/10/2013, 22:20

... the products of the first PCR before performing the second PCR, simply use a 1–5 µl aliquot of the first PCR reaction mix as the template for the second PCR Of course one potential disadvantage of ... added by PCR primers Blunt-end polish then ligate PCR products Restriction digestion of concatamers of PCR products Figure 6.1 Concatemerization of blunt-end PCR products to allow efficient restriction ... (LIC) of PCR products PCR Methods Appl 4: 172–177 Puri cation and cloning of PCR products 133 14 Haun RS, Moss J (1992) Ligation-independent cloning of glutathione S-transferase fusion genes for...

Ngày tải lên: 25/10/2013, 22:20

... degreesÆcm2Ædmol)1 by using mean residue masses of 110 RESULTS Identi cation of crucial residues for the antibacterial activity of pyrrhocoricin An Ala-scan was performed to identify key residues that could ... ordered conformation Identi cation of the pyrrhocoricin binding surface on E coli DnaK Encouraged by the success of the Ala-scan in identifying the essential residues of pyrrhororicin for antibacterial ... transport vehicles for the peptide Consistent with this, an application of the predicative algorithm for nonspecific interaction of peptides with the conventional peptide binding pocket of E coli DnaK...

Ngày tải lên: 08/03/2014, 10:20

... were screened for mutations using LapA1 genespecific primers and dideoxy-chain-termination DNA sequencing A random distribution of mutations was not obtained Therefore, several specific oligonucleotide ... studies suggested a role for waters and the bihydroxide ion in the general base mechanism of blLAP [21,24] The recent identi cation of bicarbonate in the active site of the native E coli PepA ... DYNAFIT for the analysis of enzyme kinetic data: application to HIV proteinase Anal Biochem 237, 260–273 33 Laemmli, U.K (1970) Cleavage of structural proteins during assembly of the head of bacteriophage...

Ngày tải lên: 08/03/2014, 23:20

... point for a model of the interactions of the two T cruzi proteins, which, together with our identi cation of the specificity-conferring interactions, will be important for development of drugs for ... difference of 9° between these complexes may be correlated with the variation of the rate of binding and affinity for chagasin of these enzymes On the other hand, in the two crystal forms of the chagasin–cathepsin ... structure-aided design of specific inhibitors of cruzipain and cathepsins Detailed knowledge of the structure and inhibition mode of chagasin should be valuable in guiding the development of drugs for the prevention...

Ngày tải lên: 16/03/2014, 04:20

... expressing the His-tagged forms sGas1523, sGas1E161Q and sGas1E262Q (B) Representative puri cation of sGas1523-H Silver staining of the different steps of the puri cation are shown Lane 1, culture ... were amplified using two sets of primers For sGas1E161Q-H, a pairing of the forward primer XHup and reverse primer RMGLN161 (5¢-TGTTAGTAACTTGATTACCGGCGAAG-3¢), and a pairing of the forward primer ... frozen and stored at )20 °C prior to puri cation Puri cation of His-tagged Gas1 proteins A 10–20 mL sample of the culture supernatant was dialyzed at °C for 16 h against lysis buffer (50 mM sodium...

Ngày tải lên: 16/03/2014, 18:20

... MS analysis has identified large families of isoforms of TAXI-type inhibitors in wheat grain [17] The amino acid sequences of TAXI-I and TAXI-II isoforms share a high degree of identity (UniProt ... molecular forms: an intact form with a molecular mass of approximately 40 kDa; and a processed form, consisting of two polypeptides of approximately 10 and 30 kDa, held together by one disulfide bond ... allowed identi cation of the structural determinants for the different TAXItype xylanase inhibition strengths and specificities Fine-tuned criteria could be deduced for the evaluation of TAXI-type...

Ngày tải lên: 23/03/2014, 05:22

... screening for improved variants Genetic variation was introduced by error prone polymerase chain reactions (EP -PCR) , followed by shuffling of the PCR products Cloning and transformation of the shuffling ... the twofold increased ratio of polymerization over hydrolysis at both low and high sucrose concentrations Polymerase efficiency of the improved variants The results of the iodine staining of polymer ... ‘geometric lock’ responsible for a closed conformation of a highly flexible loop in the B¢ domain Removal of the Asp231 side chain allowed simulation of large movements of this loop using geometric...

Ngày tải lên: 23/03/2014, 11:20

... levels of classi cation provide an overview of the diversity of protein structures now known and would be appropriate both for researchers and students The specific lower levels should be helpful for ... Most of the folds are assigned to one of the five structural classes on the basis of the secondary structures of which they composed: (1) all alpha (for proteins whose structure is essentially formed ... 422 537 Communication Figure In scop, the unit of classi cation is usually the protein domain Small proteins, and most of those of medium size, have a single domain and are, therefore, treated...

Ngày tải lên: 23/03/2014, 12:20

... Km for CTP formation were determined with respect to NH3 and glutamine for each of the mutant enzymes except for K187A CTPS which was inactive (Table 3) Direct examination of the conversion of ... hydrolysis of glutamine, but had a value of kcat that was half of that observed for wild-type CTPS in the absence of GTP When the concentration of GTP was increased to mM, the value of kact was ... presence of ATP Fig Time-dependent inactivation of wild-type CTPS by trypsin (A) Inactivation of CTPS-catalysed NH3-dependent CTP formation in the absence of ligands (s) and in the presence of ATP...

Ngày tải lên: 23/03/2014, 17:22

... electron micrographs of PEDOT:PSS particles, a without DMSO modi cation, b with DMSO modi cation from the films and keep the configuration unchanged, just like the heat-setting process of polymers at ... composition of the PEDOT:PSS from various depth can be analyzed [19] The S(2p) core level spectra of pristine PEDOT:PSS film and DMSO modified PEDOT:PSS film are shown in Fig 4b, c, with a take-off angle of ... Fig The size of pristine PEDOT:PSS is 100–400 nm, much larger than 30–100 nm obtained after the modi cation The PEDOT:PSS particles become much smaller and more uniform after the modi cation, thus...

Ngày tải lên: 22/06/2014, 00:20

Ngày tải lên: 26/03/2015, 10:18

... DEVELOPMENT OF PROBIOTIC PRODUCTS FOR USAGE IN SHRIMP FARMING SCHOOL OF BIOTECHNOLOGY AND FOOD TECHNOLOGY HANOI UNIVERSITY OF TECHNOLOGY DAI CO VIET, HANOI, VIETNAM Phone/Fax: +84-4 3868 2470 Email: biofotech@mail.hut.edu.vn ... Export of frozen shrimp constitutes a large proportion of the Seafood export in Vietnam However, currently the shrimpfarming in our country is facing a lot of difficulties due to the loss of shrimps ... loss of shrimps on a large scale Therefore, research on producing efficient biological products in order to improve the resistant ability of shrimps is now of great significance Currently, prawn...

Ngày tải lên: 25/05/2015, 20:00

... mechanism of ampli cation of whole cDNAs and the TSAT -PCR technique (A) Ampli cation of full-length cDNAs (B) Procedure for the nested PCR reactions This figure is reproduced from [39] Fig Schematic of ... templates for GLGI ampli cation, decreasing the complexity of the templates and thus reducing possible nonspecific ampli cation; (b) the 3¢ cDNAs are enriched by PCR to provide sufficient templates for ... used for GLGI ampli cation; and (e) direct precipitation of GLGI-amplified products and cloning into vectors without gel puri cation prevents the potential loss of amplified products These changes...

Ngày tải lên: 07/03/2014, 00:20

... Page of 11 (page number not for citation purposes) PCR positivity by the broad-range PCR amplification system PCR was positive in 20 of the 27 (74.10%) patients indicating the presence of bacterial ... extraction from synovial fluid and broad-range PCR amplification of 16S rRNA genes, cloning and sequencing of bacterial DNA For 10 minutes, 500 μl of SF were centrifuged at 10,000 g The whole SF ... samples allowing the identification of the species level Sequence analysis of the PCR- positive samples revealed the presence of a wide spectrum of bacterial DNA in SF samples of all studied patients...

Ngày tải lên: 09/08/2014, 14:22

... from Uzbekistan The PCR- RFLP-based assay yielded aberrant results for five of 54 HBV strains [7] Similar to the results reported in this study, their DNA sequencing data for two of the five strains ... PCRRFLP-based genotyping of HBV strains is simple to perform and accurately detected genotype assignment of a substantial number (60 of 80) of HBV strains Additionally, the PCR- RFLP-based method ... Comparative evaluation of INNO-LiPA HBV assay, direct DNA sequencing and subtractive PCR- RFLP for genotyping of clinical HBV isolates Virology Journal 2010, 7:111 Page of ...

Ngày tải lên: 12/08/2014, 04:20

... sec of annealing at 50 °C, and of extension at 72 °C This cycle was repeated 30 times and followed by incubation at 72 °C for 10 for the final extension The initial concentration of PCR products ... and the sequencing analysis of the PCR products with DNA extracted from activated sludge Melting curve analysis was performed after amplification The reaction mixture was heated to 95 °C for sec, ... tool for its quantification in activated sludge It would be suitable for routine monitoring or measurement of a large number of samples, and lead to better understanding of the contribution of...

Ngày tải lên: 05/09/2013, 09:38

... cleavage [41] The use of thrombin significantly increases the cost of the puri cation, and the require1276 ment for HPLC increases the length and complexity of the puri cation procedure Moreover, as with ... (Fig 2E) Puri cation of Ab(M1–40) and Ab(M1–42) The present work describes a rapid and inexpensive puri cation scheme to produce high-purity Ab(M1–40) and Ab(M1–42) in 24 h The puri cation scheme, ... studied, but electron micrographs of the edges of fibril mats or isolated well-dispersed fibers are presented to show the fibril morphology at high definition These fibrils vary in length, and can be...

Ngày tải lên: 18/02/2014, 13:20

... was amplified by PCR and purified for electroporation to O35E competent cells as described previously [19] After 24 h of incubation, the resulting Zeor colonies were selected for PCR identi cation ... NaCl ⁄ chloroform (1 : v ⁄ v) A new volume of chloroform was used for each of the three extractions Finally, the organic phase was re-extracted with de-ionized water to remove traces of salts, ... Analysis of physiochemical and biological features of both mutants was performed to study the functions of these genes and the structures of their resultant LOSs in vitro and in vivo Results Identification...

Ngày tải lên: 18/02/2014, 14:20

... HMBC NMR spectra of 17a,20,23-trihydroxyvitamin D3 are presented in the supplementary Fig S2 Identi cation of minor products of vitamin D3 metabolism by P450scc Fig Identi cation of 20,23-dihydroxyvitamin ... 40 50 Time (min) Fig Identi cation of minor metabolites of vitamin D3 Products of P450scc action on vitamin D3 with retention times of 32, 26 and 26.7 in Fig were purified, incorporated into phospholipid ... The lack of hydroxylation of vitamin D3 at position 22 now provides a clear explanation for the inability of P450scc to cleave the side chain of vitamin D3 Hydroxylation of the side chain of cholesterol...

Ngày tải lên: 18/02/2014, 17:20