Title:

Detection of DNA
To separate this DNA according the size.
electrophoresis.

Aim:
Hypothesis:

You want to detect your isolated plasmid DNA from exp 7, using

I expect the DNA quantities in samples with the length of plasmid DNA should be 3337 bp and NanoDrop ND1000 what do you mean by Nanodrop? To determine your DNA, you use a DNA ladder + control DNA.

Materials:
You use an other concentration for the control

Table 1: Overview of used hazardous compounds

Compound
name

4X Bromophenol
blue

Sybr Safe

Amount

Concentration

H/P statements

GHS H
Flammable
H226
liquid and
statemen
:
vapor
t
Keep away
from
P210 heat/sparks/ope
:
n flames/hot
surfaces. - No
smoking
Wear protective
gloves/protecti
P280
ve clothing/eye
:
protection/face
protection
GHS P
P233 Keep container
tightly closed
statemen :
Ground/Bond
t
P240 container and
:
receiving
equipment
IF ON SKIN
P303 (or hair): Take
+
off immediately
P361 all
+
contaminated
P353 clothing. Rinse
:
skin with
water/shower

Protection

Waste category

Wear protective
gloves/protective
clothing/eye
protection/face
protection.

Contain spill and

collect, as apropriate.
Transfer to a
chemical waste
container for disposal
in accordance with
local regulations.
halogenated
organic waste

Method:

Also write down the changes you made on the protocol (running time, etc.)

Results:

Image 1: Results electrophoresis

Number the lanes in your picture and show base pairs near the ladder.

From left to the right:

1.Control DNA
2.Michelle's plasmid DNA
3.Grace's plasmid DNA
4.Bianca's plasmid DNA
5.Eva's plasmid DNA
and ladder?
The pGFPuv plasmid DNA was detected on the gel and it's length was determined to be as predicted,3337 base pairs. The band that
represented linear DNA, which was compared to control DNA and marker, was very faint and in the same cases was almost undetectable
(3 and 5).
Is it really 3337 base pairs? Try to verify the base pairs from the ladder.
Discussion: Agarose gel electrophoresis tehnique was used to identify and determine the length of plasmid DNA. A tracking dye
(bromophenol blue) was added to the DNA sample to indicate the stopping point.
Agarose gel in the electrophoresis chamber was loaded with plasmid DNA and the chamber was hooked up to a power source designed
to send an electrical current through the gel. The pGFPuv plasmid DNA was detected on the gel and it's length was determined to be as
predicted,3337 base pairs. The band that represented linear DNA, which was compared to control DNA and marker, was very faint and in
the same cases was almost undetectable (3 and 5).
That was due the small concentration of plasmid DNA, although the procedure was adjusted and more DNA was added to the samples
tested on the gel. It was expected that there will be two more bands present on the gel, one below the liniar plasmid on the gel DNA band
which would represent the slower open circular DNA, and the second one above liniar DNA which would represent faster supercoiled
DNA. Those two bands would it be observed being that the concentration is very low (63.8 mg/mL). Cloudy bands were detected
underneath DNA bands, which could be a result of some RNA being present in the sample, since RNA molecules are small and able to
migrate through the gel.
Okay.

Conclusion:
The ratio of absorbance at 260 and 280 nm of the sample 4 was 1.02 compared to the 1.8 which is generally accepted as 'pure' for DNA,
which imputes that sample was impure to some extent and corresponds to the privious statement that is probably contaminated with
DNA. Show this ratio in your hypothesis as well. What size is your plasmid DNA? Could you detect it?