For transcriptional fusion do you clone the promoter of interest upstream of the reporter gene which is promoterless to study the promoter strength?
And in translational fusion do you clone the gene of interest with its promoter upstream to the promoter less reporter gene (e.g. GFP) to study the biology/chemistry of protein?
Is this correct?
As you stated, transcription fusion cloning the promoter of interest, upstream of any transcription unit, it can be reporter gene. Such construct was usually used either:
(1) to evaluate the function of the promoter (i.e. how the promoter regulate gene expression, either developmentally regulated? or inducer regulated? and where and when the transcript of the original genes are express - either spatial expression or temporal expression? Tissue specific or cell specific expression? and so on).
If you ask any of those question, then you need to conduct experiment using transcriptional fusion of the promoter with reporter gene. The use of reporter gene is to make it easier for user to evaluate and monitor when, where, and how the promoter are actively transcribing the transcript... Just follow the presence of reporter gene product and you can determine the activity of the promoter.
2) Once one knows the function of a specific promoter, then they can use the promoter to express desirable transcription unit the way the function of the promoter, by replacing reporter gene ORF with those of gene of interest.
As an example, you want to specifically express Bt Toxin only in fruits, then you will want to make transcription fusion between fruit specific promoter with Cry (Bt toxin) transcription unit plus the terminator. The chimaeric construct will express Bt toxin only in fruits in the transgenic plants.
As you also stated, the translation fusion was combining two open reading frame (one is gene of interest and the second is reporter gene such as gfp for non-destructive detection) into single one. The translation product of the fusion reading frame would be one larger polypeptides containing both domain. The gfp domain is used tag the gene of interest domain. With such translation fusion, ones could monitor where the gene of interest is expressed by monitoring where the gfp is present.
I hope these can answer some of your questions. Good luck in your research....
translational fusions are the gene of interest and a reporter gene present in the same frame under same promoter and SD sequence
As you stated, transcription fusion cloning the promoter of interest, upstream of any transcription unit, it can be reporter gene. Such construct was usually used either:
(1) to evaluate the function of the promoter (i.e. how the promoter regulate gene expression, either developmentally regulated? or inducer regulated? and where and when the transcript of the original genes are express - either spatial expression or temporal expression? Tissue specific or cell specific expression? and so on).
If you ask any of those question, then you need to conduct experiment using transcriptional fusion of the promoter with reporter gene. The use of reporter gene is to make it easier for user to evaluate and monitor when, where, and how the promoter are actively transcribing the transcript... Just follow the presence of reporter gene product and you can determine the activity of the promoter.
2) Once one knows the function of a specific promoter, then they can use the promoter to express desirable transcription unit the way the function of the promoter, by replacing reporter gene ORF with those of gene of interest.
As an example, you want to specifically express Bt Toxin only in fruits, then you will want to make transcription fusion between fruit specific promoter with Cry (Bt toxin) transcription unit plus the terminator. The chimaeric construct will express Bt toxin only in fruits in the transgenic plants.
As you also stated, the translation fusion was combining two open reading frame (one is gene of interest and the second is reporter gene such as gfp for non-destructive detection) into single one. The translation product of the fusion reading frame would be one larger polypeptides containing both domain. The gfp domain is used tag the gene of interest domain. With such translation fusion, ones could monitor where the gene of interest is expressed by monitoring where the gfp is present.
I hope these can answer some of your questions. Good luck in your research....
Thanks for the explanation
What will then be the difference then of the translation fusion and a protein tag?
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