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Transgenic sheep were first cloned in 1996 when scientists used somatic cell nuclear transfer to create Dolly the sheep. This involves removing the nucleus of an egg cell and replacing it with the nucleus of a donor adult cell, usually a fibroblast. The reprogrammed cell is then stimulated to divide and develop into an embryo. While transgenic sheep can be useful models for studying various fields like immunology and hematology, the cloning process has low success rates and cloned animals often display abnormalities. Transgenic sheep are also used commercially to produce recombinant proteins in various tissues like milk.
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Transgenic sheep ppt for staff & students
Sheep named Dolly was cloned by transfer of a nucleus from a mammary (Udder) cell of an adult sheep into an egg cell.
mammary cell
Nucleus
insert into
a egg cell
First demonstration of pluripotency (totipotency) of a nucleus of a differentiated adult cell.
Cloning of dolly somatic cell nuclei
clone cattle, sheep, goats, pigs.
nuclear transfer procedures are similar.
Adult donor cells from a variety of cell types(mammary epithelial and ovarian cells, fibroblasts, lymphocytes) are isolated
Cultured and genetically modified methods.
individual donor cells are fused to an enucleated oocyte with short-duration electric pulse.
eg: two 2.5 kilovolt /cm pulses for 10microseconds
Used to fuse adult cattle fibroblasts with enucleated oocytes.
The pulses simultaneously induce cell fusion and oocyte activation.
Blastocyst stage before transferred into the uterus of a pseudopregant female.
Confirmed transgene at the time of birth
Surviving animals produced by nuclear transfer are healthy.
There, is a substantial loss of individual before and after birth some of the cloned animals display abnormalities.
Abnormlities such as increased birth weight.
Dna methylation and histone modification of the original donor cell is inappropriate maintained in the cells of the recipient animals.
Organ culture technique in synthetic media- animal tissue culture
Organ culture is a development from tissue culture that allows for the culture of pieces of organs on artificial media to accurately model organ functions in various states. Special culture methods are required as organs require high oxygen levels. Organ pieces can be cultured on plasma clots, agar, raft methods using lens paper or rayon, grid methods, or in liquid media using supports like gauze or rafts. Organ culture faces limitations as results may not match whole animal studies due to lack of in vivo drug metabolism.
GENE TRANSFER METHODS IN ANIMALS
Gene transfer methods in animals can be natural or artificial. Natural methods include conjugation, transformation, and transduction which transfer genes between bacteria. Artificial methods like microinjection, biolistics, liposome mediated transfer, calcium phosphate mediated transfer, and electroporation are used to directly insert genes into cells. These techniques transfer genes into organisms for genetic engineering applications such as producing transgenic animals, developing vaccines, and gene therapy to treat diseases.
Lectut btn-202-ppt-l5. yeast cloning vectors (1)
Yeast cloning vectors allow DNA fragments to be replicated and expressed in yeast cells. There are several types of yeast vectors including integrating plasmids (YIps) that replicate by integrating into yeast chromosomes, episomal plasmids (YEps) that replicate independently but can also integrate, and replicating plasmids (YRps) that contain an autonomously replicating sequence (ARS) and replicate at low copy numbers. Yeast artificial chromosomes (YACs) are engineered chromosomes containing telomeric, centromeric, and ARS sequences that can clone very large DNA fragments of up to 3000 kb.
Biolistics
Biolistics, also known as gene gun technology or particle bombardment, is a physical method of DNA transfer where microprojectiles coated with DNA are accelerated into intact plant or animal cells. A gene gun works by firing microscopic gold or tungsten particles coated with DNA through a stopping screen and into target cells. Some DNA-coated particles pass through the cell wall and cytoplasm, dissociating from the particle inside the cell where the DNA may be expressed. Gene gun technology has been used successfully to transform many plant species as well as deliver DNA vaccines and label cell subsets for research.
Dna modifying enzymes
MBB 501 PLANT BIOTECHNOLOGY
INFORMATION ABOUT DIFFERENT DNA MODIFYING ENZYMES
WHAT IS AN ENZYME?
Alkaline Phosphatase
Polynucleotide kinase
Terminal deoxyneucleotidyl transferase
Nucleases
Exonuclease
Bal31 Exonuclease III
Endonuclease
S1 endonulease
Deoxyribonuclease 1 (Dnase 1)
RNase A
RNase H
Restriction Endonuclease
PvuI
PvuII
Different types of endonuclease enzymes
The recognition sequences for some of the most frequently used restriction endonucleases.
Categorization of enzymes
Isoschizomers
Neoschizomers
Isocaudomers
Microinjection
a proper description about the process microinjection and also about gene transfer. and different types of DNA delivery methods.
with advantages, disadvantages, limitations and applications.
Gene transfer in animals
This document discusses various methods for transferring genes into animal cells, including viral and non-viral approaches. Viral methods use viruses like adenovirus to transfer genes, while non-viral methods include biochemical techniques like calcium phosphate transfection, lipid-mediated transfection using lipofectamine, and physical methods like microinjection, particle bombardment/gene guns, ultrasound, and electroporation. The document provides detailed protocols for lipid-mediated transfection and some of the other non-viral methods.
Recommended
Transgenic sheep ppt for staff & students
Sheep named Dolly was cloned by transfer of a nucleus from a mammary (Udder) cell of an adult sheep into an egg cell.
mammary cell
Nucleus
insert into
a egg cell
First demonstration of pluripotency (totipotency) of a nucleus of a differentiated adult cell.
Cloning of dolly somatic cell nuclei
clone cattle, sheep, goats, pigs.
nuclear transfer procedures are similar.
Adult donor cells from a variety of cell types(mammary epithelial and ovarian cells, fibroblasts, lymphocytes) are isolated
Cultured and genetically modified methods.
individual donor cells are fused to an enucleated oocyte with short-duration electric pulse.
eg: two 2.5 kilovolt /cm pulses for 10microseconds
Used to fuse adult cattle fibroblasts with enucleated oocytes.
The pulses simultaneously induce cell fusion and oocyte activation.
Blastocyst stage before transferred into the uterus of a pseudopregant female.
Confirmed transgene at the time of birth
Surviving animals produced by nuclear transfer are healthy.
There, is a substantial loss of individual before and after birth some of the cloned animals display abnormalities.
Abnormlities such as increased birth weight.
Dna methylation and histone modification of the original donor cell is inappropriate maintained in the cells of the recipient animals.
Organ culture technique in synthetic media- animal tissue culture
Organ culture is a development from tissue culture that allows for the culture of pieces of organs on artificial media to accurately model organ functions in various states. Special culture methods are required as organs require high oxygen levels. Organ pieces can be cultured on plasma clots, agar, raft methods using lens paper or rayon, grid methods, or in liquid media using supports like gauze or rafts. Organ culture faces limitations as results may not match whole animal studies due to lack of in vivo drug metabolism.
GENE TRANSFER METHODS IN ANIMALS
Gene transfer methods in animals can be natural or artificial. Natural methods include conjugation, transformation, and transduction which transfer genes between bacteria. Artificial methods like microinjection, biolistics, liposome mediated transfer, calcium phosphate mediated transfer, and electroporation are used to directly insert genes into cells. These techniques transfer genes into organisms for genetic engineering applications such as producing transgenic animals, developing vaccines, and gene therapy to treat diseases.
Lectut btn-202-ppt-l5. yeast cloning vectors (1)
Yeast cloning vectors allow DNA fragments to be replicated and expressed in yeast cells. There are several types of yeast vectors including integrating plasmids (YIps) that replicate by integrating into yeast chromosomes, episomal plasmids (YEps) that replicate independently but can also integrate, and replicating plasmids (YRps) that contain an autonomously replicating sequence (ARS) and replicate at low copy numbers. Yeast artificial chromosomes (YACs) are engineered chromosomes containing telomeric, centromeric, and ARS sequences that can clone very large DNA fragments of up to 3000 kb.
Biolistics
Biolistics, also known as gene gun technology or particle bombardment, is a physical method of DNA transfer where microprojectiles coated with DNA are accelerated into intact plant or animal cells. A gene gun works by firing microscopic gold or tungsten particles coated with DNA through a stopping screen and into target cells. Some DNA-coated particles pass through the cell wall and cytoplasm, dissociating from the particle inside the cell where the DNA may be expressed. Gene gun technology has been used successfully to transform many plant species as well as deliver DNA vaccines and label cell subsets for research.
Dna modifying enzymes
MBB 501 PLANT BIOTECHNOLOGY
INFORMATION ABOUT DIFFERENT DNA MODIFYING ENZYMES
WHAT IS AN ENZYME?
Alkaline Phosphatase
Polynucleotide kinase
Terminal deoxyneucleotidyl transferase
Nucleases
Exonuclease
Bal31 Exonuclease III
Endonuclease
S1 endonulease
Deoxyribonuclease 1 (Dnase 1)
RNase A
RNase H
Restriction Endonuclease
PvuI
PvuII
Different types of endonuclease enzymes
The recognition sequences for some of the most frequently used restriction endonucleases.
Categorization of enzymes
Isoschizomers
Neoschizomers
Isocaudomers
Microinjection
a proper description about the process microinjection and also about gene transfer. and different types of DNA delivery methods.
with advantages, disadvantages, limitations and applications.
Gene transfer in animals
This document discusses various methods for transferring genes into animal cells, including viral and non-viral approaches. Viral methods use viruses like adenovirus to transfer genes, while non-viral methods include biochemical techniques like calcium phosphate transfection, lipid-mediated transfection using lipofectamine, and physical methods like microinjection, particle bombardment/gene guns, ultrasound, and electroporation. The document provides detailed protocols for lipid-mediated transfection and some of the other non-viral methods.
Transgenic mice
Introduction
Definition
History
Why are the transgenic animals being produced
Transgenic mice
Mice: as model organism
Methods of creation of transgenic mice
knock-out mice
Application of transgenic mice
Conclusion
References
Linker, Adaptor, Homopolymeric Tailing & Terminal Transferase
A brief idea about the problems and solutions in cloning regarding Linker, Adaptor, Homopolymeric Tailing & Terminal Transferase enzyme.
Blue white screening
This document describes the blue-white screening technique for identifying recombinant bacteria. Blue-white screening relies on the activity of β-galactosidase, an enzyme in E. coli that cleaves lactose. Plasmid vectors used in cloning carry a fragment of the lacZ gene. When the plasmid inserts foreign DNA at the multiple cloning site, it disrupts lacZ and prevents complementation, so the bacteria cannot metabolize lactose and appear white on an indicator plate. If no DNA inserts or inserts elsewhere, complementation occurs and bacteria appear blue. The technique allows rapid identification of bacteria that took up recombinant plasmid.
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Livestock sector is an important sector in indian economy. To boost the productive performance of existing livestock population in india, biotechnolgy plays a key role to fullfill this.
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Viral vectors are efficient tools for gene delivery due to viruses' ability to transfer DNA into host cells. The document discusses several types of viral vectors, including adenoviral, adeno-associated, retroviral, lentiviral, and baculovirus vectors. It provides details on the structure and genome organization of different viruses used to create these vectors. The document also explains the process of generating recombinant viral vectors by removing unnecessary viral genes and inserting genes of interest. Viral vectors allow for transient or stable gene expression and are useful for both research and clinical applications such as gene therapy and vaccine development.
MICROINJECTION GENE TRANSFER METHOD
Microinjection is a gene transfer technique where DNA is directly injected into cells using a fine glass micropipette. It is highly efficient at the individual cell level and was originally used for transfecting hard-to-transfect cells. The procedure involves holding a cell using one pipette while another pipette is used to inject DNA into the cell's cytoplasm or nucleus. It allows for stable transfection efficiencies of around 20% and is used to generate transgenic animals by injecting DNA into oocytes, eggs or embryos. However, it is time-consuming and can only be done for a small number of cells.
Bacteriophage vectors
Bacteriophage vectors
Bacteriophage
WHY BACTERIOPHAGE AS A VECTOR?
M13 phage
Genome of m13 phage
Life cycle and dna replication of m13
CONSTRUCTION M13 AS PHAGE VECTOR
M13 MP 2 vector
M13MP7 VECTOR
Selection of recombinants
Lambda replacement vectors
LAMBDA EMBL 4 VECTOR
P1 PHAGE
GENOME OF P1 PHAGE
P1 PHAGE AS VECTOR
P1 phage vector system
bacterial artificial chromosome & yeast artificial chromosome
BAC & YAC are artificially prepared chromosomes to clone DNA sequences.yeast artificial chromosome is capable of carrying upto 1000 kbp of inserted DNA sequence
Sv 40
Simian virus 40 (SV40) is a DNA virus that can cause tumors in monkeys and humans, and it was first identified as a contaminant in polio vaccines in the 1960s. SV40 has been widely used as a cloning vector due to its ability to efficiently deliver genes into a variety of cells without killing the host cell or eliciting an immune response. Future research prospects for SV40 vectors include developing recombinant versions for gene transfer applications and furthering understanding of related retroviruses.
Selection and screening of recombinant clones
This document discusses several methods for selecting recombinant clones after introducing recombinant DNA into host cells:
- Direct selection involves using a gene from the inserted DNA that confers antibiotic resistance to select clones that grow on media containing that antibiotic.
- Insertional inactivation selection works by inactivating a host gene when foreign DNA inserts into it, allowing selection of recombinants.
- Blue-white screening uses a vector with a disrupted lacZ gene; foreign DNA insertion repairs the gene, allowing recombinants to be identified by colony color.
- Colony hybridization detects recombinants by transferring colonies to a membrane and probing for the inserted DNA sequence.
- Immunological tests identify clones expressing antigens encoded by the
Transfection
The document discusses various methods of transfection in animals. Transfection is the process of introducing nucleic acids into eukaryotic cells. It describes viral transfection using bacteria like Agrobacterium tumefaciens and viruses. Non-viral methods include chemical transfection using calcium phosphate, liposomes, polyamines. Mechanical transfection employs microinjection or particle bombardment. Common chemical methods are calcium phosphate precipitation, polyplexes, and liposomes/lipoplexes. Viruses used are retroviruses, adenoviruses, adeno-associated viruses. Bacterial and viral vectors allow for integration into the host genome while chemical and mechanical are often transient.
Genomic library construction
A DNA library is a collection of DNA fragments that have been cloned into vectors. DNA libraries allow researchers to isolate and study specific DNA fragments of interest. To create a genomic library, DNA is extracted from an organism, cut into fragments, inserted into vectors, and introduced into host bacteria to generate clones containing all the organism's DNA sequences. This library can then be screened to identify and study particular genes. DNA libraries provide an efficient way to store, isolate, and analyze DNA sequences.
Phagemid vector
This document discusses the phagemid vector pBluescript, which can be used in either the positive or negative orientation. pBluescript is a phagemid vector that can be used to clone DNA fragments for sequencing, mutagenesis, protein expression or other molecular biology experiments. References for further information about pBluescript are provided.
Direct Gene Transfer Methods
This document discusses various methods of genetic transfer, including natural genetic transfer between organisms as well as technological methods developed to manipulate genes. It describes how donor DNA can enter a recipient cell and recombine, producing genetically distinct offspring. Several gene transfer technologies are then outlined, including microinjection, biolistics, calcium phosphate precipitation, lipofection, and electroporation. The document explains the basic mechanisms and applications of each method while also noting their limitations for different purposes like gene therapy. In the conclusion, it emphasizes that gene transfer technologies now allow relatively easy and accurate introduction of genes into target cells to potentially cure diseases.
Shuttle vector
1. Yeast plasmids like the 2 micron circle have been extensively studied and developed into yeast cloning vectors.
2. Shuttle vectors like YEp vectors contain selectable marker genes like LEU2 and bacterial plasmid origins of replication like pBR322, allowing them to replicate in both E. coli and yeast.
3. The 2 micron circle is a 6kb endogenous yeast plasmid that replicates autonomously through an ARS sequence and is maintained at 50-100 copies per cell.
Cell synchronization
INTRODUCTION
HISTORY
NEED OF SYNCHRONIZATION
SYNCHRONOUS CULTURES CAN BE OBTAINED IN SEVERAL WAYS:
Physical fractionation .
Chemical appro ach
CENTRIFUGAL ELUTRIATION
Inhibition of DNA synthesis
Nutritional deprivation
SYNCHRONIZATION AT LOW TEMPERATURE
CELLULAR TOTIPOTENCY
SOME HIGHLIGHTS OF CELL SYNCHRONIZATION
REFERENCES
Ti plasmid as a vector,
Introduction
Ti plasmid
Agrobacterium tumefaciens
Ti plasmid structure
Overview of infection process
Ti plasmid derived vector systems
Cointegrate vectors
Binary vectors
Agrobacterium mediated transformation of explants
Conclusions
References
Primary and established cell line culture
Introduction
Primary Culture
Steps of Primary Culture
Isolation Of Tissue
Dissection And Disaggregation
Types Of Primary Culture
Primary Explants Culture
Enzymatic Disaggregation
Mechanical Disaggregation
Cell Line( Finite & Continuous)
Naming A Cell Line
Choosing A Cell Line
Maintenance Of Cell Line
Conclusion
Reference
Primary culture and cell line
Introduction
Primary Culture
Steps In Primary Culture
Isolation Of Tissue
Dissection And/Or Disaggregation
Types Of Primary Culture
Primary Explant Culture
Enzymatic Disaggregation
Mechanical Disaggregation
Cell Line( Finite & Continuous)
Naming A Cell Line
Choosing A Cell Line
Maintenance Of Cell Line
Conclusion
reference
Yeast Artificial Chromosomes (YACs)
Yeast artificial chromosomes (YACs) are engineered DNA molecules that can clone and replicate large DNA sequences in yeast cells. YACs contain essential yeast elements like a centromere and telomeres that allow them to behave like natural yeast chromosomes. YACs can clone very large inserts of up to 10 megabases of foreign DNA, making them useful for generating whole genome libraries.
Transgenic animals and process to make transgenic animals
The document summarizes topics related to transgenic animals and gene therapy. It discusses transgenic cows, sheep, poultry, and fish. For each animal, it describes the process used to create transgenic versions, including pronuclear microinjection and somatic cell nuclear transfer. Benefits include producing human therapeutic proteins and altering milk composition. Challenges include high costs and low success rates. Gene therapy techniques like viral vectors and electroporation are explained for inserting genes into tissues to treat disease. Somatic gene therapy aims to modify individual patients while germline gene therapy alters heritable genes passed to offspring.
Cloning
Cloning involves producing genetically identical individuals through recombinant DNA technology and molecular cloning techniques. There are several types of cloning including DNA cloning, reproductive cloning, and therapeutic cloning. Reproductive cloning uses somatic cell nuclear transfer to generate an animal with the same nuclear DNA as another existing or previously existing animal. Therapeutic cloning produces human embryos for stem cell research to study development and treat diseases. While cloning research has potential benefits, there are also significant health and ethical risks involved especially for reproductive cloning in humans.
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knock-out mice
Application of transgenic mice
Conclusion
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Linker, Adaptor, Homopolymeric Tailing & Terminal Transferase
A brief idea about the problems and solutions in cloning regarding Linker, Adaptor, Homopolymeric Tailing & Terminal Transferase enzyme.
Blue white screening
This document describes the blue-white screening technique for identifying recombinant bacteria. Blue-white screening relies on the activity of β-galactosidase, an enzyme in E. coli that cleaves lactose. Plasmid vectors used in cloning carry a fragment of the lacZ gene. When the plasmid inserts foreign DNA at the multiple cloning site, it disrupts lacZ and prevents complementation, so the bacteria cannot metabolize lactose and appear white on an indicator plate. If no DNA inserts or inserts elsewhere, complementation occurs and bacteria appear blue. The technique allows rapid identification of bacteria that took up recombinant plasmid.
Biotechnology in livestock improvement
Livestock sector is an important sector in indian economy. To boost the productive performance of existing livestock population in india, biotechnolgy plays a key role to fullfill this.
Animal viral based vectors
Viral vectors are efficient tools for gene delivery due to viruses' ability to transfer DNA into host cells. The document discusses several types of viral vectors, including adenoviral, adeno-associated, retroviral, lentiviral, and baculovirus vectors. It provides details on the structure and genome organization of different viruses used to create these vectors. The document also explains the process of generating recombinant viral vectors by removing unnecessary viral genes and inserting genes of interest. Viral vectors allow for transient or stable gene expression and are useful for both research and clinical applications such as gene therapy and vaccine development.
MICROINJECTION GENE TRANSFER METHOD
Microinjection is a gene transfer technique where DNA is directly injected into cells using a fine glass micropipette. It is highly efficient at the individual cell level and was originally used for transfecting hard-to-transfect cells. The procedure involves holding a cell using one pipette while another pipette is used to inject DNA into the cell's cytoplasm or nucleus. It allows for stable transfection efficiencies of around 20% and is used to generate transgenic animals by injecting DNA into oocytes, eggs or embryos. However, it is time-consuming and can only be done for a small number of cells.
Bacteriophage vectors
Bacteriophage vectors
Bacteriophage
WHY BACTERIOPHAGE AS A VECTOR?
M13 phage
Genome of m13 phage
Life cycle and dna replication of m13
CONSTRUCTION M13 AS PHAGE VECTOR
M13 MP 2 vector
M13MP7 VECTOR
Selection of recombinants
Lambda replacement vectors
LAMBDA EMBL 4 VECTOR
P1 PHAGE
GENOME OF P1 PHAGE
P1 PHAGE AS VECTOR
P1 phage vector system
bacterial artificial chromosome & yeast artificial chromosome
BAC & YAC are artificially prepared chromosomes to clone DNA sequences.yeast artificial chromosome is capable of carrying upto 1000 kbp of inserted DNA sequence
Sv 40
Simian virus 40 (SV40) is a DNA virus that can cause tumors in monkeys and humans, and it was first identified as a contaminant in polio vaccines in the 1960s. SV40 has been widely used as a cloning vector due to its ability to efficiently deliver genes into a variety of cells without killing the host cell or eliciting an immune response. Future research prospects for SV40 vectors include developing recombinant versions for gene transfer applications and furthering understanding of related retroviruses.
Selection and screening of recombinant clones
This document discusses several methods for selecting recombinant clones after introducing recombinant DNA into host cells:
- Direct selection involves using a gene from the inserted DNA that confers antibiotic resistance to select clones that grow on media containing that antibiotic.
- Insertional inactivation selection works by inactivating a host gene when foreign DNA inserts into it, allowing selection of recombinants.
- Blue-white screening uses a vector with a disrupted lacZ gene; foreign DNA insertion repairs the gene, allowing recombinants to be identified by colony color.
- Colony hybridization detects recombinants by transferring colonies to a membrane and probing for the inserted DNA sequence.
- Immunological tests identify clones expressing antigens encoded by the
Transfection
The document discusses various methods of transfection in animals. Transfection is the process of introducing nucleic acids into eukaryotic cells. It describes viral transfection using bacteria like Agrobacterium tumefaciens and viruses. Non-viral methods include chemical transfection using calcium phosphate, liposomes, polyamines. Mechanical transfection employs microinjection or particle bombardment. Common chemical methods are calcium phosphate precipitation, polyplexes, and liposomes/lipoplexes. Viruses used are retroviruses, adenoviruses, adeno-associated viruses. Bacterial and viral vectors allow for integration into the host genome while chemical and mechanical are often transient.
Genomic library construction
A DNA library is a collection of DNA fragments that have been cloned into vectors. DNA libraries allow researchers to isolate and study specific DNA fragments of interest. To create a genomic library, DNA is extracted from an organism, cut into fragments, inserted into vectors, and introduced into host bacteria to generate clones containing all the organism's DNA sequences. This library can then be screened to identify and study particular genes. DNA libraries provide an efficient way to store, isolate, and analyze DNA sequences.
Phagemid vector
This document discusses the phagemid vector pBluescript, which can be used in either the positive or negative orientation. pBluescript is a phagemid vector that can be used to clone DNA fragments for sequencing, mutagenesis, protein expression or other molecular biology experiments. References for further information about pBluescript are provided.
Direct Gene Transfer Methods
This document discusses various methods of genetic transfer, including natural genetic transfer between organisms as well as technological methods developed to manipulate genes. It describes how donor DNA can enter a recipient cell and recombine, producing genetically distinct offspring. Several gene transfer technologies are then outlined, including microinjection, biolistics, calcium phosphate precipitation, lipofection, and electroporation. The document explains the basic mechanisms and applications of each method while also noting their limitations for different purposes like gene therapy. In the conclusion, it emphasizes that gene transfer technologies now allow relatively easy and accurate introduction of genes into target cells to potentially cure diseases.
Shuttle vector
1. Yeast plasmids like the 2 micron circle have been extensively studied and developed into yeast cloning vectors.
2. Shuttle vectors like YEp vectors contain selectable marker genes like LEU2 and bacterial plasmid origins of replication like pBR322, allowing them to replicate in both E. coli and yeast.
3. The 2 micron circle is a 6kb endogenous yeast plasmid that replicates autonomously through an ARS sequence and is maintained at 50-100 copies per cell.
Cell synchronization
INTRODUCTION
HISTORY
NEED OF SYNCHRONIZATION
SYNCHRONOUS CULTURES CAN BE OBTAINED IN SEVERAL WAYS:
Physical fractionation .
Chemical appro ach
CENTRIFUGAL ELUTRIATION
Inhibition of DNA synthesis
Nutritional deprivation
SYNCHRONIZATION AT LOW TEMPERATURE
CELLULAR TOTIPOTENCY
SOME HIGHLIGHTS OF CELL SYNCHRONIZATION
REFERENCES
Ti plasmid as a vector,
Introduction
Ti plasmid
Agrobacterium tumefaciens
Ti plasmid structure
Overview of infection process
Ti plasmid derived vector systems
Cointegrate vectors
Binary vectors
Agrobacterium mediated transformation of explants
Conclusions
References
Primary and established cell line culture
Introduction
Primary Culture
Steps of Primary Culture
Isolation Of Tissue
Dissection And Disaggregation
Types Of Primary Culture
Primary Explants Culture
Enzymatic Disaggregation
Mechanical Disaggregation
Cell Line( Finite & Continuous)
Naming A Cell Line
Choosing A Cell Line
Maintenance Of Cell Line
Conclusion
Reference
Primary culture and cell line
Introduction
Primary Culture
Steps In Primary Culture
Isolation Of Tissue
Dissection And/Or Disaggregation
Types Of Primary Culture
Primary Explant Culture
Enzymatic Disaggregation
Mechanical Disaggregation
Cell Line( Finite & Continuous)
Naming A Cell Line
Choosing A Cell Line
Maintenance Of Cell Line
Conclusion
reference
Yeast Artificial Chromosomes (YACs)
Yeast artificial chromosomes (YACs) are engineered DNA molecules that can clone and replicate large DNA sequences in yeast cells. YACs contain essential yeast elements like a centromere and telomeres that allow them to behave like natural yeast chromosomes. YACs can clone very large inserts of up to 10 megabases of foreign DNA, making them useful for generating whole genome libraries.
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Linker, Adaptor, Homopolymeric Tailing & Terminal Transferase
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Similar to transgenic-sheep-.pptx
Transgenic animals and process to make transgenic animals
The document summarizes topics related to transgenic animals and gene therapy. It discusses transgenic cows, sheep, poultry, and fish. For each animal, it describes the process used to create transgenic versions, including pronuclear microinjection and somatic cell nuclear transfer. Benefits include producing human therapeutic proteins and altering milk composition. Challenges include high costs and low success rates. Gene therapy techniques like viral vectors and electroporation are explained for inserting genes into tissues to treat disease. Somatic gene therapy aims to modify individual patients while germline gene therapy alters heritable genes passed to offspring.
Cloning
Cloning involves producing genetically identical individuals through recombinant DNA technology and molecular cloning techniques. There are several types of cloning including DNA cloning, reproductive cloning, and therapeutic cloning. Reproductive cloning uses somatic cell nuclear transfer to generate an animal with the same nuclear DNA as another existing or previously existing animal. Therapeutic cloning produces human embryos for stem cell research to study development and treat diseases. While cloning research has potential benefits, there are also significant health and ethical risks involved especially for reproductive cloning in humans.
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Transgenic animal prof.a.k.saha
Transgenic animals are genetically engineered to contain extra genes. This seminar discusses methods for creating transgenic animals like microinjection, embryonic stem cell transfer, and nuclear transfer. Microinjection involves injecting foreign genes into fertilized eggs. Embryonic stem cell transfer modifies stem cells in culture before implanting in blastocysts. Nuclear transfer fuses donor cell nuclei with egg cells. Examples given include producing human proteins in cow's milk and human hemoglobin in pigs. While transgenic animals could benefit humans, the process is inefficient and risky for animal health.
Cloning
This document discusses various aspects of artificial insemination and cloning in livestock. It defines cloning as DNA cloning, reproductive cloning, and therapeutic cloning. It then covers topics like semen collection, ovum collection, embryo collection, artificial insemination procedures, the use of AI in different livestock industries, embryo splitting, cryopreservation of genetic material, estrus synchronization, and more.
TRANSGEIC ANIMALS MICE BIRD.pptx
It's include all the details about the transgenic technology.all the techniques like micro injection,SCNT,pro nuclear injection method.It include all the Transgenic mice bird and fish.
genetically modified organisms
Genetically modified organisms (GMOs) are organisms whose genetic material has been altered using genetic engineering techniques. Common GMOs include crop plants and microbes. The genetic material is typically transferred between different species, which would not occur naturally. This allows for precise and fast introduction of traits compared to traditional breeding methods. While GMOs hold promise for increasing crop yields and producing medicines, there are also health and environmental safety concerns that require further research.
Applications of Biotechnology in Agriculture for Water Security, Nguyen Van T...
Biotechnology applications in agriculture can help improve water security. Advances in embryo technology, cloning, and transgenic animals can lead to smarter livestock farming practices. This includes using techniques like ICSI and embryo transfer to rescue rare and infertile animals or produce livestock with desirable traits. Somatic cell nuclear transfer can also be used to conserve species. Additional biotechnologies such as induced pluripotent stem cells may allow the production of human organs in animals through knockout of specific genes, helping regenerative biomedicine and pharmaceutical production. Overall, smart applications of biotechnology in agriculture can enhance sustainable development and water security.
Extract the following requirements from the Research below Articl.pdf
Extract the following requirements from the Research below:
Article Interspecific Nuclear Transfer Blastocysts Reconstructed from Arabian Oryx Somatic
Cells and Domestic Cow Ooplasm Aiman A. Ammari *Q, Muath G. ALGhadi G, Ramzi A.
Amran, Nawal M. Al Malahi and Ahmad R. Alhimaidi Department of Zoology, College of
Science, King Saud University, PO. Box 2455, Riyadh 11451, Saudi Arabia * Correspondence:
alammari@ksuedu.si Simple Summary: Interspecies SCNT-based cloning and in vitro
production of interspecies SCNTderived embryos allows cells that have undergone terminal
differentiation to be reprogrammed to become totipotent cells. The development of the
interspecific Somatic cell nuclear transfer oryx in vitro for the first time in Saudi Arabia will
promote additional research on animal reproductive doning for the preservation of endangered
species. Abstract: Cloning, commonly referred to as somatic cell nuclear transfer (SCNT), is the
technique of enucleating an oocyte and injecting a somatic cell into it. This study was carried out
with interspecific SCNT technology to clone the Arabian Oryx utilizing the oryx's fibroblast
cells and transfer it to the enucleated oocytes of a domestic cow. The recipient oocytes were
extracted from the cows that had been butchered. Oryx somatic nuclei were introduced into cow
oocytes to produce embryonic cells. The study was conducted on three groups, Oryx
interspecific somatic cell nuclear transfer into enucleated cocytes of domestic cows, cow SCNT
"the same bovine family species", used as a control group, and in vitro fertilized (IVF) cows to
verify all media used in this work. The rates of different embryo developmental stages varied
slightly (from 1-cell to morula stage). Additionally, the oryx check for interspecies Somatic cell
nuclear transfer blastocyst developmental rate (9.23%) was comparable to that of cow SCNT
(8.33\%). While the blastula stage rate of the (IVF) cow embryos exhibited a higher Citation:
Ammari, A.A; cleavage rate (42%) in the embryo development stage. The results of this study
enhanced domestic ALGhadi, M.G: Amran, RA: cow oocytes' ability to support interspecific
SCNT cloned oryx, and generate a viable embryo that Al Malahi, N.M.: Alhimaidi, A.R. can
advance to the blastula stage. Interspecific Nuclear Transfer Blastrcysts Reourstructed fixm
Arakian Cryx Sombtic Cells and Keywords: iSCNT; cloning; Arabian Oryx; blastocyst Domestic
Cow Ooplasm. Vet. Sc. 2023, 10, 17. https://doiorg/ 10.3590/ vetsi100110017 1. Introduction
Academic Editors: Pengiki Lin, By the early 1970s, the Arabian Oryx had become extinct in the
wild but was still Dong Zhwa and Junwei Li thriving in zoos and private preserves. The Arabian
Oryx was the first animal to be desigReccived: 30 Nonember 2002 nated as vulnerable in 2011
after being listed as extinct in the wild and subsequently as Revised: 22 December 2122
endangered in 1986 on the International Union for Conservation of Nature's (IUCN) Red
Acceptrat: 26 De.
Animal cloning
This document discusses animal cloning, specifically somatic cell nuclear transfer (SCNT). It provides information on the history of cloning, animals that have been cloned, the SCNT process, challenges to successful cloning including reprogramming differentiated cells, and problems seen in cloned animals including embryonic and postnatal abnormalities. Applications of cloning such as restoring endangered species, generating transgenic animals, and gene knockout in farm animals are also covered.
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Extract the following requirements from the Research below Articl.pdf
Extract the following requirements from the Research below Articl.pdf
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Syllabus
Chapter-1
Introduction to objective, scope and outcome the subject
Chapter 2
Introduction: Scope and Specialization of Civil Engineering, Role of civil Engineer in Society, Impact of infrastructural development on economy of country.
Chapter 3
Surveying: Object Principles & Types of Surveying; Site Plans, Plans & Maps; Scales & Unit of different Measurements.
Linear Measurements: Instruments used. Linear Measurement by Tape, Ranging out Survey Lines and overcoming Obstructions; Measurements on sloping ground; Tape corrections, conventional symbols. Angular Measurements: Instruments used; Introduction to Compass Surveying, Bearings and Longitude & Latitude of a Line, Introduction to total station.
Levelling: Instrument used Object of levelling, Methods of levelling in brief, and Contour maps.
Chapter 4
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Chapter 5
Transportation: Introduction to Transportation Engineering; Traffic and Road Safety: Types and Characteristics of Various Modes of Transportation; Various Road Traffic Signs, Causes of Accidents and Road Safety Measures.
Chapter 6
Environmental Engineering: Environmental Pollution, Environmental Acts and Regulations, Functional Concepts of Ecology, Basics of Species, Biodiversity, Ecosystem, Hydrological Cycle; Chemical Cycles: Carbon, Nitrogen & Phosphorus; Energy Flow in Ecosystems.
Water Pollution: Water Quality standards, Introduction to Treatment & Disposal of Waste Water. Reuse and Saving of Water, Rain Water Harvesting. Solid Waste Management: Classification of Solid Waste, Collection, Transportation and Disposal of Solid. Recycling of Solid Waste: Energy Recovery, Sanitary Landfill, On-Site Sanitation. Air & Noise Pollution: Primary and Secondary air pollutants, Harmful effects of Air Pollution, Control of Air Pollution. . Noise Pollution Harmful Effects of noise pollution, control of noise pollution, Global warming & Climate Change, Ozone depletion, Greenhouse effect
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2. Satheesh Gopi, Basic Civil Engineering, Pearson Publishers.
3. Ketki Rangwala Dalal, Essentials of Civil Engineering, Charotar Publishing House.
4. BCP, Surveying volume 1
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transgenic-sheep-.pptx
- 1. Biotechnology Transgenic sheep Iv – semester PRESENTED BY SIVARAMAKRISHNAN.T II M.SC. MICROBIOLOGY Reg.no.20201232516121 PRESENTED TO DR.G.RAMANATHAN GANESHAN DEPARTMENT OF MICROBIOLOGY SRI PARAMAKALYANI COLLEGE ALWARKURICHI
- 2. Transgenic sheep 1996-DOLLY-the sheep was cloned using cells from a 6yr old sheep's udder by somatic cell nuclear transfer (SCNT)technology somatic cell nuclear transfer technology involves two different cells ovum somatic cell (eggoocytes) (skin fibroblast) -nucleus microinjected with nucleus transgenic DNA discarded cell enucleated egg discarded (empty ovum) lone somatic nucleus
- 8. E.g.; inactivating both alleles of a locus or creating multiple gene changes Individual donor cells are fused to an enucleated oocyte with short-duration electric pulse E.g.; two 2.5 kilovolt /cm pulse for 10microseconds Used to fuse adult cattle fibroblasts with enucleated oocytes The pulse simultaneously indce cell fusion and oocyte activation Blastocyst stage before transferred into the uterus of a pseudopregant female
- 10. Surviving animals produced by nuclear transfer are healthy There is a substantial loss of individual before and after birth some of the cloned animals display abnormalities Abnormliaties such as increased birth weight DNA methylation and histone modification of the original donor cell is inappropriate maintained in the cells of the recipient animal
- 11. USES OF TRANSGENIC SHEEP it is used as a model for studying Immunology Human Blood Clotting Factor Viii Transplantation Haematology Biological Product Manufacturing Recombinant DNA Drug Production In Milk
- 12. Advantages Site-specific Genetic Changes Are Possible All Offspring Are Transgenic In DNA Microinjection
- 13. Transgene Integration Occurs At Random Sites Unstable Things Are Formed Expression Is Always Constrained No.Of Generations Required Much Devoted Id Needed Nucleic From Somatic Cell Is Used Develop Perfect Cloning Of Livestock
- 14. DISADVANTAGES Difficult Procedure Failed In Vitro Fertilization Expensive
- 15. Industrial biotechnology and commodity products transgenic animals are being used for production of recombinant proteins in milk, egg, white, blood, urine seminal plasma ,and silk worm cocoons The amount milk produced by animals (L yr-1) are 8000 per cow, 1000 per goat, 300 per sheep, and per rabbit