DNA QuantificationContinuation of Lab 4

• Qiagen DNAeasy kit. • Quantify our yields using a Nanodrop system.

DNeasy Minispin Column

Collection tube

Eppendorph

Figure 4.Summary of DNA extraction protocol.II.A. DNA Extraction and Purification

II. DNA Extraction, Purification and Quantification

1. Use pipet to transfer DNA mixture into DNeasy minispin column

4. Save column.

2. Centrifuge at 8000 rpm for 1 min.3. Discard liquid and collection tube.

STEP 5

1. Use new collection tube2. Add 500 µl of AW1 buffer3. Centrifuge 1 minute at 8000 rpm4. Discard flow-through and collection tube5. Keep column

Flow-through

Collection tube

STEP 6

1. Use new collection tube2. Add 500 µl of AW2 buffer3. Centrifuge 3 minute at 14000 rpm4. Discard flow-through and collection tube5. Keep column

Flow-through

Collection tube

STEP 7

STEP 8

1. Place Dneasy mini-spin column in 1.5 ml microcentrifuge tube (eppindorph)

2. Snip cap off of column

microcentrifuge tube

STEP 91. Add 100 µl of AE buffer over disk in column2. Incubate at room temp for 1 minute3. Centrifuge 1 minute 8000 rpm4. DO NOT DISCARD FLOW-THROUGH

disk

STEP 10

1. Label with sharpie at top of lid

2. Freeze and save for PCR Lab 10

1. Add another 100 µl of AE buffer over disk in column and repeat procedure below

2. Incubate at room temp for 1 minute3. Centrifuge 1 minute 8000 rpm4. DO NOT DISCARD FLOW-THROUGH

Nanodrop spectrometer used to measure DNA concentrations

II.B. DNA Quantitation