Expression of N-terminal seven amino acids peptide of fibrin (β–peptide) on surface of M13KO7...
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بسم الله الرحمن الرحیم
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Expression of N-terminal seven amino acids peptide of
fibrin (β–peptide) on surface of M13KO7 phage
Supervisor: Dr. Hossein KhanahmadAref Farrokhi [email protected]
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Introduction
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Vascular accidents
• Acute myocardial infarction
• Cerebrovascular accident (CVA) or acute cerebral infarction
• Pulmonary thromboembolism
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Fibrinolytic drugs for therapy
Urokinase (UK) Streptokinase (SK) Recombinant Tissue plasminogen activator
(rTPA)
Side effects:
• Low efficiency • Hemorrhage
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Clot targeting for preventing
side effects and elevating the
fibrinolysis efficiency Local enrichment using fibrin targeting : Specific anti fibrin antibody(MA-15C5), 59D8): Chemical conjugation Fusion protein(Ab-E)
Nanobody
Scfv FabWhole
Ab
Fv fragmen
t
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Material and methods
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fibrinogen beta chain isoform 1 preproprotein
[Homo sapiens]
Beta-peptide as an antigen
Gly-His-Arg-Pro-Leu-Asp-LysFSARGHRPLDKKREE
45-52
NCBI Reference Sequence: NP_005132.2
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Bam HI
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BsrG I
10211119 bp
7550 bp
GTCTGTACACCGTTCATCTGTCC
TGGATCCTCATTAAAGCCAGAA
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1301..1522
1579..2853
gene="V" CDS 843..1106
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After overlap PCR
1. Agarose gel electrophoresis of PCR product2. Gel extraction3. Ligation between ABC fragment and plasmid
backbone
1179 bp
8729 bp7550 bp
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After transformation to top10
Plasmid extraction from transformed top10 bacteria
Transformation of phage genome to TG1
PCR on recombinant phage genome
Confirmation by sequencing
Obtaining phages from culture for SDS PAGE
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SDS PAGE
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Thank you for your attention
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