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Dna Extraction Of Blood And Analysis Of Phosphate Acquisition Mutants Of E. Coli Assignment

727 words - 3 pages

EXPERIMENT 1: Quantitation of DNA Solutions Using Absorbance at 260 nm

Q1.
0.137

0.279

7
13

Graph 1; DNA Absorbance at 260 nm- Standard curve depicting theoretical A260 and of 1/5 and 1/10 diluted DNA A260

An average A260 value of 0.279 for 1/5 diluted DNA was obtained, which correlated to the CDNA of 13 ?g/mL. Similarly, an average A260 value of 0.137 for 1/10 diluted DNA was obtained, which correlated to the CDNA of 7 ?g/mL. The two value of CDNA obtained differed by 6 ?g/mL.

Q2.
DNA STOCK CONCENTRATION

SAMPLE
A260
A260
A260 MEAN
CDNA DILUTION
CDNA STOCK

1/5 DILUTED DNA
0.262
0.296
0.279
13
65

1/10 DILUTED DNA
0.131
0.143
0.137
7
70

Table 1; DNA Stock Concentration
CALCULATION OF CDNA STOCK:

For 1/5 Diluted DNA: For 1/10 Diluted DNA:
= (5) dilution factor X CDNA dilution = (10) dilution factor X CDNA dilution
5 X 13= 65 ?g/mL 10 X 7= 70 ?g/mL

The CDNA stock solution of 65 ?g/mL obtained from 1/5 diluted DNA, and the CDNA stock solution of 70 ?g/mL obtained from 1/10 diluted DNA, statistically only differed by 5 ?g/mL. The values for both CDNA stock solutions are statistically insignificant, due to minor difference in values.

Q3.

Concentration DNA Stock: 68.783 Mg/mL Volume Stock: 1 mL
Concentration Blood Solution: 1:1 Volume Blood Solution: 7 mL

mL of Blood = 3.5 mL

Mg DNA= 68.783 Mg in 3.5 mL blood

Concentration DNA in blood= 68.783/3.5
= 19.652 Mg/mL
DNA YIELD= 19.652 Mg/mL

According to a study conducted by S, Ghatak, R, Muthukumaran & S Nachimuthu (2013), a concentration of 57?94 ng/?l of DNA in blood (from 50 ?l) samples was obtained. The results based on the study, differs significantly from the obtained DNA concentration of 19.652 Mg/mL. Similarly, the product manual suggests that an isolated DNA of high purity (A260/A260 ratio:1.7-1.9) will yield 20-35 ?g/mL, which only varies by 0.348 ?g/mL from the obtained value of 19.652 Mg/mL (Jayachandran, A 2013).

EXPERIMENT 2: ANALYSIS OF PHOSPHATE ACQUISITION MUTANTS OF E. COLI

Q1.
ANALYSIS OF PHOSPHATE ACQUISITION MUTANTS OF E. COLI

STRAIN
GROWTH ON Pi PLATES
GROWTH ON G3P PLATES
AP-ASE ASSAY COLOUR

1
NO
YES
YELLOW

...

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