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Reporter gene and gene fusions T.Y.Bsc

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pratik prajapati

In this PPT You can learn briefly about Reporter Gene and Gene fusion And Gene manipulation method. Reference From Microbiology. Author - Brock. 12th Edition,

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? The Mandavi Education Society T.Y.Bsc(Microbiology) Krishna prajapati Teaching Assistance(Microbiology)
Reporter gene and gene fusions
Introduction ? DNA manipulation has revolutionary the study of gene regulation. ? A coading sequence from one source(the reporter)may be fused to a regulatory region from another source. ? Such gene fusions are often used in studying gene regulation especially where assaying the level of the natural gene product is difficult or time consuming. ? They may also be used to increase expression of desired gene product.
Reporter genes ? The key property of reporter gene is that it encodes a protein that is easy to detect and assay. ? Reporter gene are used for variety of purposes. ? They may used to report on the presence or absence of particular genetic elements(such as plasmid)or DNA inserted within the vector. ? They can also fused to other genes or to promoter of other genes so that gene expression can be studied.
LacZ gene ? The first gene to be used widely as reporter gene was lacZ, which encodes the enzyme ¦Â-galactosidase. ? Cells expressing ¦Â- galactosidasecan be detected by easily by their color on indicater plates that contain the artificial substrates Xgal ,which cleaved by ¦Â-galactosidase to yield blue colour.
Blue white screening .
Conti..
Green flourescent protein(GFP) ? Another widely used reporter gene encodes luciferase. ? This enzymes makes cells expressing it luminescent. colonies containing the reporter system can be detected by on agar plate by their luminscense against a large background of other colonies.
Conti.. ? However,the expression of luciferase depends on more than one gene beacause several accesary factors are needed. ? By contrast, the green fluroscent protein needs no accessary factors and is widely used as reporter ? Gene for was GFP was originally cloned from the jelly Aequorea victoria,the GFP protein may be expressed in most bacterial cell. ? It is stable and causes little or no disruption of host cell metabolism.
Conti¡­ ? If expression of cloned gene linked to that of GFP, expression of GFP signals that the cloned gene has also been expressed. ? Recent advance in fluorescent labeling now allow the simultaneous use of multiple fluorescent markers.
Some like that..
Gene fusion.. ? It is possible to engineering construct that consist of segment from two different genes. ? such construct are known as gene fusions.
Conti.. ? If the promoter that controls a coding sequence is removed, the coding sequence can be fused to different regularly region to place the gene under the control of different promoter alternatively. ? The promoter region can be fused to gene whose product is east to assay.
There are two types of gene fusion Fig.. . Operon fusion ? In operon fusion, a coading sequence that retain its own translation start site and signals is fused to the transcriptional signals of another gene.
Gene fusion Protein fusion ? In protein fusion, two coading sequence are fused with the result that they share the same transcriptional and traslational start sites and signals. Fig¡­
Conti¡­. ? Gene fusion are often used in studying gene regulation, especially if measuring the level of natural gene product is difficult and time consuming. ? The reulatery region of the gene of interest is fused to coding sequence for reporter gene, such as ¦Â- galactosidase or GFP. ? The reporter is then made under the condition that would trigger expression of the target gene. ? The expression of reporter is assay under variety of condition to determine how the gene of interest is regulated, where as protein fusion reveal translational cantrol.
Reporter construction
Conti.. ? Gene fusion may also be used to test for the effect of regulatory genes. ? Mutation that effect regulatory genes are inroducted into cells carrying gene fusion, and expression is measured and compared to cell lacking the regulatory mutation. ? This allows the rapid screening of multiple regulatory genes that are suspected of controlling the target gene.
Reporter gene and gene fusions T.Y.Bsc

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Reporter gene and gene fusions T.Y.Bsc

  • 1. ? The Mandavi Education Society T.Y.Bsc(Microbiology) Krishna prajapati Teaching Assistance(Microbiology)
  • 2. Reporter gene and gene fusions
  • 3. Introduction ? DNA manipulation has revolutionary the study of gene regulation. ? A coading sequence from one source(the reporter)may be fused to a regulatory region from another source. ? Such gene fusions are often used in studying gene regulation especially where assaying the level of the natural gene product is difficult or time consuming. ? They may also be used to increase expression of desired gene product.
  • 4. Reporter genes ? The key property of reporter gene is that it encodes a protein that is easy to detect and assay. ? Reporter gene are used for variety of purposes. ? They may used to report on the presence or absence of particular genetic elements(such as plasmid)or DNA inserted within the vector. ? They can also fused to other genes or to promoter of other genes so that gene expression can be studied.
  • 5. LacZ gene ? The first gene to be used widely as reporter gene was lacZ, which encodes the enzyme ¦Â-galactosidase. ? Cells expressing ¦Â- galactosidasecan be detected by easily by their color on indicater plates that contain the artificial substrates Xgal ,which cleaved by ¦Â-galactosidase to yield blue colour.
  • 8. Green flourescent protein(GFP) ? Another widely used reporter gene encodes luciferase. ? This enzymes makes cells expressing it luminescent. colonies containing the reporter system can be detected by on agar plate by their luminscense against a large background of other colonies.
  • 9. Conti.. ? However,the expression of luciferase depends on more than one gene beacause several accesary factors are needed. ? By contrast, the green fluroscent protein needs no accessary factors and is widely used as reporter ? Gene for was GFP was originally cloned from the jelly Aequorea victoria,the GFP protein may be expressed in most bacterial cell. ? It is stable and causes little or no disruption of host cell metabolism.
  • 10. Conti¡­ ? If expression of cloned gene linked to that of GFP, expression of GFP signals that the cloned gene has also been expressed. ? Recent advance in fluorescent labeling now allow the simultaneous use of multiple fluorescent markers.
  • 12. Gene fusion.. ? It is possible to engineering construct that consist of segment from two different genes. ? such construct are known as gene fusions.
  • 13. Conti.. ? If the promoter that controls a coding sequence is removed, the coding sequence can be fused to different regularly region to place the gene under the control of different promoter alternatively. ? The promoter region can be fused to gene whose product is east to assay.
  • 14. There are two types of gene fusion Fig.. . Operon fusion ? In operon fusion, a coading sequence that retain its own translation start site and signals is fused to the transcriptional signals of another gene.
  • 15. Gene fusion Protein fusion ? In protein fusion, two coading sequence are fused with the result that they share the same transcriptional and traslational start sites and signals. Fig¡­
  • 16. Conti¡­. ? Gene fusion are often used in studying gene regulation, especially if measuring the level of natural gene product is difficult and time consuming. ? The reulatery region of the gene of interest is fused to coding sequence for reporter gene, such as ¦Â- galactosidase or GFP. ? The reporter is then made under the condition that would trigger expression of the target gene. ? The expression of reporter is assay under variety of condition to determine how the gene of interest is regulated, where as protein fusion reveal translational cantrol.
  • 18. Conti.. ? Gene fusion may also be used to test for the effect of regulatory genes. ? Mutation that effect regulatory genes are inroducted into cells carrying gene fusion, and expression is measured and compared to cell lacking the regulatory mutation. ? This allows the rapid screening of multiple regulatory genes that are suspected of controlling the target gene.