Kunming Institute of Zoology
Question
Asked 3 August 2021
Why the expression of gene was normal in cloned individual although the transgenic vector was inserted successfully?
Hello, everyone. I used a transgenic vector for SCNT and successfully obtained cloned individuals. I screened the cloned individuals for the presence of vector by PCR and distributed individuals into 2 groups: Group 1 containing transgenic vector and Group 2: without transgenic vector (called control). Next, I performed QPCR to evaluate the expression of gene. I found individuals with transgenic vectors (Group 1) also had overexpression of gene. But there were also few individuals in group 1 that although contains a transgenic vector but the expression of the specific gene was similar to the control group (without vector). So, need an expert opinion that why the few individuals who presented normal gene expression was with transgenic vector?
Thanking you in Advance.
Kind Regards
Most recent answer
No, that's not a mistake and I think S. Sudarsono answered well. Moreover, there could be random insertion
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All Answers (4)
IPB University
Dear Muhammad Ameen Jamal:
You said that you have transgenic plants, i.e. integrating transgene in the plant genome. However, there was no transgene expression.
In such case, the absence of expression might be due to many different causes. Among the causes were as follow (there should probably be more reasons):
(1) The integrated transgene may be incomplete, i.e. part of the integrated transgene is missing (either lack of the promoter, lack (part) of the coding sequence or lack of terminator. There have been reports indicating broken or incomplete transgene may actually be integrated into the genome, which eventually causing no expression of the integrated transgene.
(2) The transgene may have been integrated in the heterochromatin region (Positional effects) which results in no transgene expression in the plant tissues. Because of the transgene integration position is in the heterochromatin region, the inserted contruct was not expressed.
(3) The integrated transgene was not expressed because of silencing phenomena. In this case, even if the transgene was integrated in the euchromatin (therefore there is no positional effect), no expression was observed because of the gene silencing.
Which of those were the real cause of no expression need further studies (supporting data) before ones can conclude the reasons.
Hopefully these explanations help you figure out what were going on in your experiment. Good luck
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Kunming Institute of Zoology
Bundle of Thanks S. Sudarsono .
Sirius University of Science and Technology
Dear Mr. Muhammad Ameen Jamal,
did you make qRT-PCR ones or several times from different cDNA?
I mean, can it be the thechnical mistake?
Kunming Institute of Zoology
No, that's not a mistake and I think S. Sudarsono answered well. Moreover, there could be random insertion
1 Recommendation
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