Purifying DNA to Estimate its Purity using PCR Amplification of VNTR to Load and Run Agarose Gel
Introduction
The study of this experiment was the Dopamine transporter gene. This gene is associated with different brain disorders like bipolar, as well as certain behavioural traits such as ADHD.[1] Dopamine transporter gene is a presynaptic plasma protein containing different VNTRs in it’s UTR and plays an important role in restricting the activity of dopamine by rapid reuptake into the presynaptic neuron. DAT is part of Na+ and Cl- dependent family with the addition of other neurotransmitter transporters such as, GABA, serotonin, glycine, and norepinephrine transporters. The 3' UTR of this gene contains a 40 bp tandem repeat.[4] Dopamine …show more content…
As well as DNA, RNA molecules are also able to absorb UV light at 260nm as they contain amino acids, therefore, they both contributed to the total result.
Agarose Gel Electrophoresis of DAT VNTR PCR Products
DNA denatures causing a smear as when the gene is stuck to an enzyme, the DNA fragments can be seen in a smear on an agarose gel after performing electrophoresis.[5] For this test, a small sample of DNA that had been isolated was loaded into a well in the agarose gel which was then exposed to an electric field. DNA is negatively charged so moved towards the anode. Due to small fragments of DNA moving faster, the DNA was separated in size. This can be seen in figure 1.
IMG_0698.JPG
Figure 1
Image of agarose gel taken by a UV transilluminator.
A few PCR bands can be seen in figure 1. …show more content…
However, some affecting factors were unavoidable and can be classed as human error. In future, these small mistakes can be avoided by just simply working at a steady pace and ensuring all equipment is working correctly such as the timer on the powerpack. Furthermore, practice should be done for pipetting samples into the agarose gel so that there is no hesitation, overfilling or underfilling during the procedure. If these slight errors were not to contribute to the experiment next time, I am certain the results will portray clear PCR bands for a more detailed
Introduction
The study of this experiment was the Dopamine transporter gene. This gene is associated with different brain disorders like bipolar, as well as certain behavioural traits such as ADHD.[1] Dopamine transporter gene is a presynaptic plasma protein containing different VNTRs in it’s UTR and plays an important role in restricting the activity of dopamine by rapid reuptake into the presynaptic neuron. DAT is part of Na+ and Cl- dependent family with the addition of other neurotransmitter transporters such as, GABA, serotonin, glycine, and norepinephrine transporters. The 3' UTR of this gene contains a 40 bp tandem repeat.[4] Dopamine …show more content…
As well as DNA, RNA molecules are also able to absorb UV light at 260nm as they contain amino acids, therefore, they both contributed to the total result.
Agarose Gel Electrophoresis of DAT VNTR PCR Products
DNA denatures causing a smear as when the gene is stuck to an enzyme, the DNA fragments can be seen in a smear on an agarose gel after performing electrophoresis.[5] For this test, a small sample of DNA that had been isolated was loaded into a well in the agarose gel which was then exposed to an electric field. DNA is negatively charged so moved towards the anode. Due to small fragments of DNA moving faster, the DNA was separated in size. This can be seen in figure 1.
IMG_0698.JPG
Figure 1
Image of agarose gel taken by a UV transilluminator.
A few PCR bands can be seen in figure 1. …show more content…
However, some affecting factors were unavoidable and can be classed as human error. In future, these small mistakes can be avoided by just simply working at a steady pace and ensuring all equipment is working correctly such as the timer on the powerpack. Furthermore, practice should be done for pipetting samples into the agarose gel so that there is no hesitation, overfilling or underfilling during the procedure. If these slight errors were not to contribute to the experiment next time, I am certain the results will portray clear PCR bands for a more detailed