Making viral dna_ii
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1. Viruses must replicate their genomes to produce new virus particles. This requires the expression of viral proteins after infection. 2. DNA replication is initiated at specific origin sites using RNA primers and proceeds in the 5' to 3' direction. Host cell proteins also contribute to viral DNA replication. 3. Large DNA viruses encode most of their own replication machinery, while smaller viruses utilize host cell proteins and orchestrate the replication process through viral proteins.
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DNA replication is the process by which a cell makes an identical copy of its DNA. It involves unwinding the double helix at an origin of replication and using each parental strand as a template to synthesize new daughter strands. This results in two identical copies of the DNA molecule. Replication is semi-conservative, meaning each new DNA molecule contains one original parental strand and one newly synthesized strand. It is also bidirectional and semi-discontinuous. The leading strand is copied continuously while the lagging strand is copied discontinuously in fragments that are later joined.
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Viruses are obligate intracellular parasites that contain either DNA or RNA. They replicate through a series of steps within a host cell. There are two main viral life cycles: lytic and lysogenic. The lytic cycle involves virus replication, assembly, and lysis of the host cell. The lysogenic cycle involves integration of the viral genome into the host cell genome without immediate cell lysis. Viruses are also classified based on their genome type and replication strategy, such as retroviruses which contain RNA and replicate through a DNA intermediate.
Dna replication eukaryotes
DNA replication is the process by which DNA copies itself in living cells. It occurs in three main steps: initiation, elongation, and termination. Initiation begins at origins of replication, where proteins assemble into pre-replication complexes. During elongation, helicase unwinds the DNA strands and DNA polymerase adds complementary nucleotides to each strand. Termination occurs when the replication forks meet, with telomerase ensuring complete replication of chromosome ends.
DNA Replication in eukaryotes and prokaryotes
1. DNA replication is the process by which daughter DNA molecules are synthesized from a parental DNA template. It ensures the genetic information is transferred to the next generation with high fidelity.
2. Replication occurs semi-conservatively such that each new double helix contains one strand from the original parent DNA and one newly synthesized strand. It also occurs bidirectionally from an origin of replication.
3. DNA polymerases are the key enzymes that catalyze DNA synthesis. Other important enzymes and proteins include primase, helicase, topoisomerase, ligase, and single-stranded DNA binding proteins. Together they facilitate the initiation, elongation and termination of DNA replication.
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This document provides an overview of recombinant DNA technology. It discusses the key discoveries that led to the development of this technology, such as Watson and Crick's discovery of DNA structure. It describes the goals and basic procedure of recombinant DNA technology, including isolating DNA, cutting DNA with restriction enzymes, joining DNA together, and amplifying the recombinant DNA in bacteria. It also discusses the enzymes, vectors like plasmids and phage, and techniques used in this technology, as well as its applications in fields like agriculture, medicine, and pharmacology.
RECOMBINENT DNA.pptx
The document presents information on recombinant DNA technology. It begins with an introduction explaining that recombinant DNA is artificially created by combining DNA sequences from different species. It then discusses some of the key tools and steps used in recombinant DNA technology, including restriction enzymes that cut DNA at specific sequences, DNA ligase that joins DNA fragments, vectors like plasmids and phages that allow DNA fragments to replicate, and suitable host cells like bacteria and yeast. The document provides examples of commonly used vectors and restriction enzymes and briefly outlines the basic process of recombinant DNA technology.
DOC-20230508-WA0012..pptx
The document presents information on recombinant DNA technology. It begins with an introduction to recombinant DNA, which involves combining DNA sequences from different species. It then discusses some of the key developments in recombinant DNA technology, including Paul Berg creating the first hybrid DNA molecule in 1972 and Herman Boyer and Stanley Cohen developing techniques to clone genes in 1973. The document also outlines the basic principles and steps of recombinant DNA technology, including isolating a gene of interest, inserting it into a vector, introducing the vector into a host cell, and harvesting the gene product. It describes some important tools used in recombinant DNA, such as restriction enzymes, ligase, vectors, and suitable host cells.
Recombinent DNA
This document provides an overview of recombinant DNA technology. It discusses the discovery of DNA structure in 1953 which led to the development of this technology. It describes the goals and basic procedure, including isolating DNA, cutting DNA with restriction enzymes, joining DNA fragments, and amplifying the recombinant DNA in bacterial hosts. It also discusses the enzymes, vectors like plasmids and phages, techniques such as PCR and gel electrophoresis, and applications in agriculture, medicine, and other fields.
Replication.pptx
Viruses replicate within host cells through a series of steps: attachment, penetration, uncoating, replication of viral genes/genome, assembly, and release. A one-step growth curve experiments infects cells at the same time to study how virus count increases over time, with an eclipse period between infection and release followed by a rise period where new virus is released. Key events during replication include binding to cell receptors, energy-dependent penetration, uncoating of the virus, transcription and translation of viral genes, replication of the viral genome, assembly of new viral particles, and release through exocytosis or cell lysis.
Recombinant-DNA-upkj2r.pptx
Recombinant DNA is DNA formed by combining DNA from multiple sources. It is created through molecular cloning techniques. The process involves isolating the desired DNA fragment, inserting it into a vector DNA, introducing the vector into a host cell, and allowing the host cell to replicate, generating multiple copies of the recombinant DNA. Examples of applications include producing insulin by introducing the human insulin gene into bacteria, gene mapping, disease research, and gene therapy.
Recombinant-DNA-upkj2r.pptx
Recombinant DNA is DNA formed by combining DNA from multiple sources. It is created through molecular cloning techniques. The process involves isolating the desired DNA fragment, inserting it into a vector DNA, introducing the vector into a host cell, and allowing the host cell to replicate, generating multiple copies of the recombinant DNA. Examples of applications include producing insulin by introducing the human insulin gene into bacteria, mapping genes, and developing genetically modified organisms.
Biotecnología
1) Gene cloning involves inserting foreign DNA into plasmids which are then inserted into bacterial cells. The bacterial cells rapidly multiply, producing multiple copies of the gene of interest.
2) Restriction enzymes and DNA ligase are used to cut and join DNA fragments to create recombinant DNA. Probes are used to screen libraries and identify clones containing genes of interest.
3) Cloned genes can be expressed in bacterial or eukaryotic cells to produce large quantities of the gene's protein product. This allows researchers to study protein function.
Viruses
Viruses replicate by taking over the host cell's machinery. They produce mRNA using their genome as a template. This mRNA directs the production of viral proteins, including structural proteins that make up the capsid and non-structural proteins that help with replication. The virus growth cycle involves adsorption to receptors, entry, uncoating, synthesis of new viral components, assembly, and release. DNA viruses replicate in the nucleus while RNA viruses replicate in the cytoplasm, except retroviruses and influenza which enter the nucleus. New viruses are assembled and released to infect other cells.
Adenovirus as an animal vector
Viruses can be used to deliver genetic material into target cells. Viruses are composed of genetic material encapsulated in a protein coat. They inject their DNA into target cells, and the viral DNA can be altered to contain a gene of interest. This allows the gene of interest to be delivered into the target cell without producing new viral particles. Adenoviruses are non-enveloped DNA viruses that can infect both dividing and non-dividing cells. They are used as vectors for gene delivery by deleting early genes and adding the gene of interest.
Genetics chapter 7 dna structure and replication
The document summarizes key discoveries related to DNA as the genetic material. It describes early evidence that DNA is the molecule responsible for heredity, including Griffith's experiments showing transformation of bacteria with heat-killed DNA and Avery, MacLeod, and McCarty's experiments showing DNA is the transforming principle. It also summarizes Hershey and Chase's experiments demonstrating that DNA enters the host bacterial cell during viral infection. The document then covers Watson and Crick's discovery of the double helix structure of DNA and key features like base pairing and the sugar-phosphate backbone. It also summarizes semiconservative replication of DNA and key molecules involved like DNA polymerase, primers, and proofreading. Finally, it discusses telomer
Cloning and recombinant dna
Cloning produces genetically identical copies of organisms. Cloning of plants and animals advanced genetic technology by improving crops and domestic animals. In the 1950s, Charles Steward cloned carrots by growing single carrot cells into full plants. Animals can be cloned through embryo splitting or cell fusion, which produced Dolly the sheep. DNA cloning is important for finding and transferring genes between species, and for applications like gene therapy. Recombinant DNA technology links DNA fragments to self-replicating vectors to clone DNA in host cells like E. coli. The polymerase chain reaction (PCR) amplifies specific DNA segments and copies DNA faster than conventional cloning methods without enzymes, vectors or host cells.
Cloning
Cloning involves producing genetically identical copies of biological material such as DNA, cells, or whole organisms. The main steps of DNA cloning are: 1) cutting the DNA fragment to be cloned and the vector DNA with the same restriction enzymes, 2) joining the fragment to the vector via ligation, 3) introducing the recombinant DNA into host cells via transformation, 4) selecting transformed cells using antibiotic resistance markers on the vector, and 5) screening clones to identify those containing the inserted DNA fragment. Common cloning vectors include plasmids, which are small extra-chromosomal DNA molecules that replicate within host bacteria or yeast cells.
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Making viral dna_ii
- 1. 1
- 2. 2
- 3. 3
- 4. DNA Replication • Viruses must replicate their genomes to make new progeny • This always requires expression of at least one virus protein, sometimes many (hence always delayed after infection) • DNA is always synthesized 5’ – 3’ • Replication initiates at a defined origin (Ori) using a primer • The host provides other proteins 4
- 5. The 5’ end problem elongate 3’ 5’ 5’ 3’ RNA primers DNA template 3’ 5’ 5’ 3’ 3’ 5’ 5’ 3’ excise primers, elongate, ligate Now what? 5
- 6. Two Basic Modes of Replication 6
- 7. Where Does the DNA Polymerase Come From? • Small DNA viruses do not encode an entire genome replication system -encode proteins that orchestrate the host -Papillomaviridae, Polyomaviridae, Parvoviridae • Large DNA viruses encode most of their own replication systems -Herpesviridae, Adenoviridae, Poxviridae 7
- 8. 8
- 9. Protein Priming ITRs Single-stranded DNA Template Displaced ss Template DNA 9
- 10. Lessons from bacteriophage ϕ29 10
- 12. Replication of adenovirus genome • An example of strand displacement synthesis • Utilizes a protein primer • Origin is at both ends • DNA polymerase is viral • Other viral proteins involved: terminal protein, ssDNA binding protein • Viral early proteins (E1a) induce quiescent cells into S phase 12
- 13. Adenovirus DNA replication requires cellular proteins 13
- 14. 14
- 15. Herpes simplex virus • HSV has 2 identical oriS and a unique oriL that is active in terminally differentiated neurons – role in transition from latent to productive infection? • DNA enters as linear molecule converts to circle • Replicates as a rolling circle 15
- 16. 16
- 17. Viral proteins for Herpes simplex virus genome replication • UL5, 8 and 52 - form primase, helicase • UL42 - a processivity protein • UL9 - Origin Binding Protein • UL29 - ssDNA Binding Protein • UL30 - DNA polymerase • 5 enzymes of nucleic acid metabolism, such as TK • Necessary but not sufficient! 17
- 18. Poxviruses • All viruses discussed replicate in nucleus • Poxvirus - cytoplasmic factories - DNA synthesis is independent of cellular proteins 18
- 19. Vaccinia DNA factories DNA stained blue; viral DNA binding protein (I3) stained red DNA only I3 only merge 19
- 20. Vaccinia DNA factories DNA stained blue; viral DNA polymerase (E9) stained red 20
- 21. Poxvirus DNA replication From “Fundamentals of MolecularVirology” by Nicholas H. Acheson, John Wiley & sons, 2007 21
- 22. Poxvirus DNA replication enzymes 22
- 23. Delayed synthesis of virion structural proteins prevents • When new viral DNAs are encapsidated they cannot be used as templates for additional DNA synthesis • Consequently genome packaging into particles is delayed – transcriptional control 23
- 24. Inhibition of cellular DNA synthesis • When viral DNA replication is carried out mostly by viral proteins, cell DNA synthesis is often inhibited • Increases availability of substrates • Adenovirus, herpesvirus, poxvirus • Mechanisms not understood 24
- 25. Viral DNAs are synthesized in specialized intracellular 25
- 26. Mechanism of exponential viral DNA replication Cellular retinoblastoma (rb) gene: Rb protein controls entry into S from G1 Loss is associated with tumors = tumor suppressor gene 26
- 27. Adenovirus E1A protein binds Rb 27
- 28. Limited replication of viral DNA • Most DNA viruses: exponential replication of genomes • Some establish long-term relationships with cells, number of genomes is limited • Various mechanisms effect genome copy number 28
- 29. Rep78/68 a site & strand- specific endonuclease Parvovirus DNA Replication 29
- 30. Parvovirus DNA replication • DNA replicates only in cells coinfected with helper adenovirus • Adenoviral helper proteins allow synthesis of large quantities of Rep 78/68 30
- 31. Parvovirus DNA replication • When no helper adenovirus is present, Rep 78/68 level is low • Little viral DNA synthesis occurs • Genome integrates into host cell DNA 31
- 32. Papillomaviruses: Controlled and exponential replication from a single Ori 32