Gene expression profile analysis dna microarrays and pitfalls
NAME- ADITYA WADHWA
2021
QUESTION 1-
· flowering is seasonal since these plants flower only in spring and flowering lasts just a few weeks.
As a result, it is not easy to collect enough biomass from petals every year for the commercial production of X.
You are the lucky postdoc who has to run this project. You need to design a strategy to conduct this project.
You need to:
5. Expression of genes X in leaves could change the expression profiles of other genes. How can you assess the global molecular changes in leaf cells? : 20 marks
Your report:
Our company is growing plants that produces an important chemical X in the petals of flower and important genes responsible for this are Gene XA, Gene XB and Gene XC. Genes are expressed and regulated by the help of TFX factor which is an important transcriptional factor. Unfortunately, this plant is not having a advantage because its commercial role is very less so cannot be used for commercial purpose, as it is not giving many flowers, flowers are small and they are seasonal based also. Thus enough biomass cannot be collected every year having a disadvantage. So, in order to increase the production of our company, we decided to re-direct its production of this important chemical X from flowers to leaves of that particular same plant. It would give us more advantages like- the size of leaves will be bigger, the growth of leaves will not depends on season and not only genome will be same in leaves and flowers but also important genes would be existing there in that particular leaves.
Flowchart diagram 1-
No problem of particular seasonal growth, and
All three genes will be there.
| REGULATORY GENES | STRUCTURAL GENES |
|---|
The promoter region is located at 5’ upstream region of a gene. Promoter plays a great role in gene expression, as switching on and off the genes. In the process of DNA transcription, transcriptional factor recognizes the promoter sequences and then binds with the region of promoter and gave the responsibility to RNA polymerase and here DNA transcription is started. Promoters of three types namely, constitutive, tissue and inducible promoters [3]. Some examples like SCR, SRK and NtHSP3A in plant transformation basically. [4] Enhancers are short 50 to 1500 base pairs regions bounded by activator proteins. Another reporter genes gets attached to regulatory sequences to find the location of expressed proteins basically [5].
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As the transcriptional factor here, TFX which is controlling and regulating the all the three genes namely, Gene XA, Gene XB and Gene XC which is from plants to flower, so we have to make an important strategy that we need to first silence these three genes first of all by using gene silencing mechanism methods like- RNA interference (RNAi) and antisense oligonucleotides (ASOs). From these methods of gene silencing we would be silencing these genes. Now, the next strategy will be that we will be constructing a transgene which is very important and that will be re-direct the production of chemical X from flowers to leaves (here we are basically changing the path of chemical X ) and after that this foreign gene or new or transgene needs to be inserted in some particular vector and for plants - the tumor-inducing or Ti plasmid can be used because our vector will be working here as a carrier which will carry our foreign gene or a transgene. Then after that the next strategy will be that we will be inserting this vector having a transgene into our old parent plant and then new expression will be there from this new transgene and then our main application major work producing chemical X will be started again with more advantages like-
Big leaves will be there,
Vector mediated gne transfer -
Agrobacterium –mediated gene transfer= Its advantage is that it is more effective in wide range of plants. [9]
These are various techniques. [10]
ANSWER 4-
Expression of genes X in leaves could change the expression profiles of other genes. We can assess the global molecular changes in leaf cells by-
The first technique is Southern Blotting which is a molecular method used for detecting the specific DNA sequences within samples of DNA. It is also used to detect how many transgenes are inserted into a parent genome and after this integrity and rearrangement can be also done. [12] and [13].
Lee-Yoon Low, Shun-Kai Yang, De-Xian Andrew Kok, Janna Ong-Abdullah, Ngai-Paing Tan and Kok-Song Lai (September 19th 2018). Transgenic Plants: Gene Constructs, Vector and Transformation Method, New Visions in Plant Science, Özge Çelik, IntechOpen, DOI: 10.5772/intechopen.79369. Available from: https://www.intechopen.com/chapters/63134
Debnath M, Prasad GBKS, Bisen PS. Molecular Diagnostics: Promises and Possibilities. Dordrecht: Springer Netherlands; 2010
Barker RF, Idler KB, Thompson DV, Kemp JD. Nucleotide sequence of the T-DNA region from the Agrobacterium tumefaciens octopine Ti plasmid pTi15955. Plant Molecular Bio-logy. Nov 1983;2(6):335-350
Van Montagu M, Zambryski P. Agrobacterium and Ti plasmids. In: Brenner’s Ency-clopedia of Genetics. Amsterdam: Elsevier; 2013. pp. 55-57
Gheysen G, Montagu MV, Zambryski P. Integration of Agrobacterium tumefaciens transfer DNA (T-DNA) involves rearrangements of target plant DNA sequences. Pro-ceedings of the National Academy of Sciences of the United States of America. Sep 1987;84(17):6169-6173
Bubner B, Baldwin IT. Use of real-time PCR for determining copy number and zygosity in transgenic plants. Plant Cell Reports. Nov 2004;23(5):263-271
