| 1. |
- Stahl, Stefan, et al.
(author)
-
Recombinant expression of insulin C-peptide
- 1997
-
Patent (pop. science, debate, etc.)abstract
- The invention relates to a method for the preparation of insulin C-peptide which includes expressing in a host cell of a multimer polypeptide containing copies of the insulin C-peptide, and degradation of the expressed polypeptide for the release of individual copies of the insulin C-peptide. The invention also relates to molecules of the nucleinic acid, to vectors of expression and host cells for the application of this method and to multimer insulin C-polypeptide expressed and degraded in accordance with the method. 21 claims
|
|
| 2. |
- Gräslund, Torbjörn, et al.
(author)
-
Production of a Thermostable DNA Polymerase by Site-Specific Cleavage of a Heat-Eluted Affinity Fusion Protein
- 1997
-
In: Protein Expression and Purification. - : Elsevier BV. - 1046-5928 .- 1096-0279. ; 9, s. 125-132
-
Journal article (peer-reviewed)abstract
- A novel strategy is described for bacterial expression and affinity purification of a recombinant truncated version of the heat-stable DNA polymerase I fromThermus aquaticus.The DNA polymerase ([Delta]Taq) was produced as a fusion to a serum albumin binding affinity handle (ABP) derived from streptococcal protein G. Based on the thermostability of the [Delta]TaqDNA polymerase, affinity-purified ABP-[Delta]Taqcould be heat-eluted from HSA columns by incubation at 85ï¿œC. To produce free [Delta]TaqDNA polymerase, efficient site-specific cleavage of the affinity tag was performed using a recombinant coxsackievirus 3C protease (3Cpro), also produced as an ABP affinity fusion. Thus, an integrated strategy could be devised where both the cleaved ABP affinity tag and the protease fusion could be recovered after site-specific cleavage using HSA-affinity chromatography. The flow-through fraction contained essentially pure [Delta]TaqDNA polymerase with full enzymatic activity.
|
|
| 3. |
- Ljungqvist, Charlotta, et al.
(author)
-
Derivatives of the B or Z domain from staphylococcal protein A (SPA) interacting with at least one domain of human factor VIII
- 1999
-
Patent (pop. science, debate, etc.)abstract
- The present invention relates to modified polypeptides which are derivatives of the B domain or Z domain from staphylococcal protein A (SPA), wherein between 1 and 20 amino acid residues of the said B or Z domain have been substituted by other amino acid residues, said substitution being made without substantial loss of the basic structure and stability of the said B or Z domain, and said substitution resulting in interaction capacity of the said polypeptide with at least one domain of human Factor VIII protein.
|
|
| 4. |
- Ljungqvist, Charlotta, et al.
(author)
-
Receptor structures
- 1999
-
Patent (pop. science, debate, etc.)abstract
- The present invention relates to modified polypeptide derivatives of the B domain or Z domain of staphylococcal protein A (SPA). The derivatives contain amino acid substitutions that result in the ability of the B domain or Z domain to interact with at least one domain of a human Factor VIII protein, without substantially disrupting the structure and stability of the B domain or Z domain.
|
|
| 5. |
- Nilsson, Peter, et al.
(author)
-
Detection of mutations in PCR products from clinical samples by surface plasmon resonance
- 1997
-
In: Journal of Molecular Recognition. - 0952-3499 .- 1099-1352. ; 10:1, s. 7-17
-
Journal article (peer-reviewed)abstract
- Two different strategies for scanning and screening of mutations in polymerase chain reaction (PCR) products by hybridization analysis are described, employing real-time biospecific interaction analysis (BIA) for detection, Real-time BIA was used to detect differences in hybridization responses between PCR products and different 17-mer oligonucleotide probes, For the analysis using a biosensor instrument, two different experimental formats were investigated based on immobilization of either biotinylated PCR products or oligonucleotide probes onto a sensor chip, Applied on the human tumour suppressor p53 gene, differences in hybridization levels for full-match and mismatch situations employing both formals allowed the detection of point mutations in exon 6 PCR products, derived from a breast tumour biopsy sample, In addition, a mutant sample sequence could be detected in a 50/50 background of wild type exon 6 sequence, The suitability of the different formats for obtaining a regenerable system and a high throughput of samples is discussed.
|
|
| 6. |
- Nilsson, Peter, et al.
(author)
-
Mutational scanning of PCR products by subtractive oligonucleotide hybridization analysis
- 1999
-
In: BioTechniques. - : Future Science Ltd. - 0736-6205 .- 1940-9818. ; 26:2, s. 308-
-
Journal article (peer-reviewed)abstract
- Here, we describe a new approach for mutational scanning of PCR products through hybridization analysis between complementary oligonucleotides. Sets of overlapping probe oligonucleotides complementary to wild-type (WT) sequence are hybridized to microbead-immobilized PCR products under solution-like conditions. Mismatch-hybridization situations between a mutant sample and probe oligonucleotides result in higher remaining concentrations in solution of involved probe oligonucleotides. Post-hybridization supernatants are subsequently analyzed for their probe oligonucleotide compositions using surface plasmon resonance-based biosensor technology Relative remaining probe oligonucleotide concentrations are monitored in real-time through hybridization analysis between probe oligonucleotides and their corresponding sensor-chip immobilized complementary counterparts. This allows for the construction of composition diagrams revealing the existence and approximate location of a mutation within an investigated sample DNA sequence. Applied on PCR products derived from clinical samples of microdissected tumor biopsies, single mutations in exons 6 and 7 of the human p53 tumor-suppressor gene were successfully detected and approximately localized.
|
|
| 7. |
- Nilsson, Peter, et al.
(author)
-
Quantitative Investigation of the Modular Primer Effect for DNA and Peptide Nucleic Acid Hexamers
- 1999
-
In: Analytical Biochemistry. - : Elsevier BV. - 0003-2697 .- 1096-0309. ; 269:1, s. 155-161
-
Journal article (peer-reviewed)abstract
- The effect on oligonucleotide–template duplex stability upon cohybridization of adjacently annealing oligonucleotides, the modular primer effect, was studied with biosensor technology. DNA and peptide nucleic acid (PNA) hexamer modules and sensor chip-immobilized template DNA strands were designed for analysis of nick, overlap, and gap modular hybridization situations. The fast hybridization kinetics for such hexamer modules allowed for the determination of apparent duplex affinities from equilibrium responses. The results showed that the hybridizational stability of modular hexamer pairs is strongly dependent on the positioning, concentration, and inherent affinity of the adjacently annealing hexamer module. Up to 80-fold increases in apparent affinities could be observed for adjacent modular oligonucleotide pairs compared to affinities determined for single hexamer oligonucleotide hybridizations. Interestingly, also for coinjections of different module combinations where DNA hexamer modules were replaced by their PNA counterparts, a modular primer effect was observed. The introduction of a single base gap between two hexamer modules significantly reduced the stabilization effect, whereas a gap of two bases resulted in a complete loss of the effect. The results suggest that the described biosensor-based methodology should be useful for the selection of appropriate modules and working concentrations for use in different modular hybridization applications.
|
|
| 8. |
- NILSSON, PETER, et al.
(author)
-
REAL-TIME MONITORING OF DNA MANIPULATIONS USING BIOSENSOR TECHNOLOGY
- 1995
-
In: Analytical Biochemistry. - : Elsevier BV. - 0003-2697 .- 1096-0309. ; 224:1, s. 400-408
-
Journal article (peer-reviewed)abstract
- The potential of real-time biospecific interaction analysis technology for applications in molecular biology is described. DNA fragments are immobilized onto a biosensor surface using the high-affinity streptavidin-biotin system and subsequently used to monitor different unit operations in molecular biology, e.g., DNA strand separation, DNA hybridization kinetics, and enzymatic modifications. A model system comprising six oligonucleotides was used, which can be assembled into a 69-bp double-stranded DNA fragment. Using this system, the biosensor approach was employed to analyze multistep solid-phase gene assembly and the performance of different enzymes routinely used for the synthesis and manipulation of DNA. In addition, a concept for the determination of single-point mutations in DNA samples is described.
|
|
| 9. |
- Nygren, Per-Åke, et al.
(author)
-
In vitro selection and optional identification of polypeptides using solid support carriers
- 1999
-
Patent (pop. science, debate, etc.)abstract
- The present invention relates to a nucleic acid molecule or a polypeptide that is selected from a method that includes cell-free expression of nucleic acid molecules immobilized on a sold support system. The present invention also relates to a molecular library that has a solid support system having a plurality of nucleic acid molecules immobilized thereon.
|
|
| 10. |
- O'Meara, Deirdre, et al.
(author)
-
Capture of single-stranded DNA assisted by oligonucleotide modules
- 1998
-
In: Analytical Biochemistry. - : Elsevier BV. - 0003-2697 .- 1096-0309. ; 255:2, s. 195-203
-
Journal article (peer-reviewed)abstract
- Real-time biospecific interaction analysis was employed to monitor direct capture of a hepatitis C virus (HCV) derived polymerase chain reaction (PCR) product by nucleic acid hybridization. Different formats for hybridization were used to study the interaction between a single-stranded HCV PCR product and capture oligonucleotides immobilized on a sensor chip via streptavidin-biotin chemistry. By employing a prehybridization step in solution with nonbiotin oligonucleotides complementary to the single-stranded target and adjacent to the immobilized probe, a significant capture was achieved in comparison to the low capture efficiency obtained using single immobilized probes (9-36 mer). High capture efficiencies were also observed when shorter immobilized probes were used in combination with strings of adjacently positioned prehybridized probes (i.e., modules). Interestingly, the introduction of single nucleotide gaps between prehybridized and/or immobilized probes dramatically reduced the capture efficiency. These results suggest that flexible systems for capture could be designed from libraries of short oligonucleotides (9 mers) used in module fashion, taking advantage of stacking interactions between the oligonucleotides. The potential applications of such oligonucleotide-assisted capture systems are discussed.
|
|
