c. Match base-pair sizes of HindIII fragments with bands that appear in the idea

Question

c. Match base-pair sizes of HindIII fragments with bands that appear in the ideal digest. Label each band with kilobase-pair (kbp) size. For example, 23,130 bp equals 23.1 kbp.

d. Set up semilog graph paper with distance migrated as the x (arithmetic) axis and base-pair length as the y (logarithmic) axis. Then, plot distance migrated versus base-pair length for each HindIII fragment.
e. Connect data points with a best-fit line.
f. Locate on x-axis the distance migrated by the first EcoRI fragment. Using a ruler, draw a vertical line from this point to its intersection with the best-fit data line.
g. Now extend a horizontal line from this point to the y-axis. This gives the calculated base-pair size of this EcoRI fragment.
h. Repeat Steps f and g for each EcoRI and BamHI fragment. Enter results in the calculated base pairs (Cal. bp) columns for each digest

Explanation / Answer

Extrapolations from the graph are most accurate for short to mid-size DNA fragments and least accurate for very large fragments. The 0.8% gel used in this experiment most effectively separates short to mid-size fragments.

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