Hello Darragh, great question. Since the peaks are expression profiles during conjugation in the mating process of meiosis, differing heights will not have an effect on ability to repair. There is a vast array of functions that Hyproc14 could be playing in the cell and this profile does not reflect its ability to perform a cellular function.
Good question Morgan, The Ftt18 gene has introns that are included in the PCR product when using genomic DNA as a template. Correct me if I am wrong, but I am assuming you are referencing it to the cDNA template. The cDNA template is DNA that is produced through reverse transcriptase of the mRNA, which will not include introns. Because the cDNA is strictly exons, there will be a shorter product. However, if you were referencing Hyproc14 or Rad51, the difference is strictly because they are genes of differing base size, this does not have significance in the experiment.
Very good question Charlotte! cDNA is coding DNA that is produced through the process of reverse transcriptase using mRNA as a template. It ultimately converts the mRNA into DNA, this process was done because it proportionally indicates the expression levels of the gene and it is more stable to quantify than mRNA directly. It is important to know that it includes only exons, as the splicing of the mRNA has already occurred. Contrarily, gDNA is the genomic DNA and does not show patterns of expression levels, this is directly from the genome of the organism and includes all introns.
Good question Alyssa! In terms of Hyproc14, little to no research has been performed on this gene. There are some other researchers in the lab that performed the same research on Hyproc14 and found results that were in consensus to my conclusion. However, DNA damage repair is extensively studied and can reveal important information such as how to get triple negative breast cancer cells to be targeted for cellular elimination. I encourage you to look into where other research is being applied, as it is incredibly vast.
how would the measuring protein levels be beneficial to the experiment – specifically, will it improve data on DNA damage, or is it used for something else?
Great question Farida! Hyproc14 may have expression that is not directly indicative of protein levels. The protein may persist (longer half-life) and mRNA be degraded quite rapidly. We see the potential of quantifying protein levels to just further indicate any differences in the gene’s products. Hope this helps!
Because you know that Rod51 is involved in gene repair, and there have been multiple genes from this class shown to likely not be involved in gene repair, do you think you would be able to compare the gene sequences of genes like Hyproc14 to Rod51 to determine which sequences are important for a gene to be able to repair DNA?
Very good question Erin, I do think this is a good thought, however the DNA damage repair (DDR) pathway is complex and there are numerous proteins playing different roles in the repair pathway. We did look for Hyproc14 homology in differing organisms and did not find any significant hits. I suppose this process may work to find genes that are similar to Rad51 in that they are performing the exact same specific function in DNA damage repair. The goal of our research was to find a gene involved in the DDR pathway and not specifically playing the same role as Rad51. Hope this helps!
Good question Abigail, we believe that the northern blot test will allows us to find more conclusive results. This test will allow for direct measurement of mRNA levels, without going through the process of reverse transcriptase PCR. It is possible this could provide better qualitative analysis. The western blot test will allow us to know exact protein level changes. We may not find anything surprising with these experiments, but it would help to further support our conclusion.
Do you think that the first peak of the Hyproc14 gene being shorter than Rad51 affected its ability to repair?
Hello Darragh, great question. Since the peaks are expression profiles during conjugation in the mating process of meiosis, differing heights will not have an effect on ability to repair. There is a vast array of functions that Hyproc14 could be playing in the cell and this profile does not reflect its ability to perform a cellular function.
Is there any significance behind the Ftt18 gDNA having about 1500bp compared to the other gene?
Good question Morgan, The Ftt18 gene has introns that are included in the PCR product when using genomic DNA as a template. Correct me if I am wrong, but I am assuming you are referencing it to the cDNA template. The cDNA template is DNA that is produced through reverse transcriptase of the mRNA, which will not include introns. Because the cDNA is strictly exons, there will be a shorter product. However, if you were referencing Hyproc14 or Rad51, the difference is strictly because they are genes of differing base size, this does not have significance in the experiment.
What is the difference between cDNA and gDNA?
Very good question Charlotte! cDNA is coding DNA that is produced through the process of reverse transcriptase using mRNA as a template. It ultimately converts the mRNA into DNA, this process was done because it proportionally indicates the expression levels of the gene and it is more stable to quantify than mRNA directly. It is important to know that it includes only exons, as the splicing of the mRNA has already occurred. Contrarily, gDNA is the genomic DNA and does not show patterns of expression levels, this is directly from the genome of the organism and includes all introns.
What other research being done by scientists can be applied, or related, to your conclusions?
Good question Alyssa! In terms of Hyproc14, little to no research has been performed on this gene. There are some other researchers in the lab that performed the same research on Hyproc14 and found results that were in consensus to my conclusion. However, DNA damage repair is extensively studied and can reveal important information such as how to get triple negative breast cancer cells to be targeted for cellular elimination. I encourage you to look into where other research is being applied, as it is incredibly vast.
how would the measuring protein levels be beneficial to the experiment – specifically, will it improve data on DNA damage, or is it used for something else?
Great question Farida! Hyproc14 may have expression that is not directly indicative of protein levels. The protein may persist (longer half-life) and mRNA be degraded quite rapidly. We see the potential of quantifying protein levels to just further indicate any differences in the gene’s products. Hope this helps!
Because you know that Rod51 is involved in gene repair, and there have been multiple genes from this class shown to likely not be involved in gene repair, do you think you would be able to compare the gene sequences of genes like Hyproc14 to Rod51 to determine which sequences are important for a gene to be able to repair DNA?
Very good question Erin, I do think this is a good thought, however the DNA damage repair (DDR) pathway is complex and there are numerous proteins playing different roles in the repair pathway. We did look for Hyproc14 homology in differing organisms and did not find any significant hits. I suppose this process may work to find genes that are similar to Rad51 in that they are performing the exact same specific function in DNA damage repair. The goal of our research was to find a gene involved in the DDR pathway and not specifically playing the same role as Rad51. Hope this helps!
What benefits do you think the blot tests will provide to your results?
Good question Abigail, we believe that the northern blot test will allows us to find more conclusive results. This test will allow for direct measurement of mRNA levels, without going through the process of reverse transcriptase PCR. It is possible this could provide better qualitative analysis. The western blot test will allow us to know exact protein level changes. We may not find anything surprising with these experiments, but it would help to further support our conclusion.