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Beta galactosidase

This is a full lab report, including sections of Abstract, Introduction, Materials &
Methods, Results, Discussion (or combined Results & Discussion) and References.
Your report should follow the format described on introductory page xi and xii of the lab
manual. Please also include detailed calculations, which may be hand-written if you
prefer and presented as an appendix.
General Comments:
Your lab report should be in the style of a scientific research paper. As such, you should
write your report so that any scientist with general knowledge of biochemical techniques
can understand it. When you present your data/results, it should be clear what the
purpose of each experiment was, what was being measured/detected in each case, and
how this data led you to your conclusions. Remember that you are telling your reader a
story, so you need to guide them through the logic of what you did, why you did it, and
what you discovered. As a story, tell of the isolation, purification and partial
characterization of β-galactosidase from E. coli. Do not present experiments as
separate units as though they are not part of an overall story.
• The Abstract should be a stand-alone paragraph that summarizes the main findings
and conclusions of your study.
• For the Introduction section, it is useful to give some background on what E. coli β-
galactosidase is and why it is of interest to study. There are many papers already
posted on Glow that will help you get this information.
• For the Materials and Methods section, you may reference the laboratory manual for
the bulk of the procedures. However, things like dilution factors for enzyme assays,
volume of Bugbuster used for cell lysis, bed volume of the nickel affinity column, etc,
that were not specified in the laboratory manual need to be indicated in this section.
• Include in the Results section all of the data that you used to generate conclusions in
your report. Be sure to provide text to put the figures and tables in context for the
reader.
• For the Discussion section, it is always interesting to compare your results to
information from the published literature.
CHEM/BIOL/BIMO 321 Lab Report 2
• Any outside sources used as well as the laboratory manual should be referenced.
Use a proper citation such as Kaplan, L. J. and Gehring, A., “Biochemistry I: Structure
and Function of Biological Molecules, Laboratory Manual” 2015, p. 27.
Specific Data Analysis Comments:
• For β-galactosidase purification: Indicate how you decided which fractions to pool to
obtain the pure β-galactosidase sample. Calculate the specific activity of both the crude
β-galactosidase (BugBuster cell lysate) and the purified β-galactosidase (pooled elution
fractions). What fold-purification of β-galactosidase was achieved as a result of the
purification procedure (specific activity pure / specific activity crude)?
• For SDS-PAGE of purification fractions: Use the gel to comment on each step of the
purification process. Measure the distance that each molecular weight standard
migrated from the top of the gel. Plot the log of the standards’ molecular weight versus
the distance migrated to generate a standard curve. Estimate the molecular weight of
the purified β-galactosidase from this standard curve. (The molecular weights of the
standards are given in the lab manual or are on Glow.)
• Comment on the success of the β-galactosidase purification (from both the actual
purification procedure and the gel observations).
• β-galactosidase enzyme kinetics: Prepare a Michaelis-Menten graph (direct plot) and
a Lineweaver-Burk graph (double reciprocal plot) of the velocity data at different ONPG
concentrations. Use these to calculate Vmax and Km for β-galactosidase. Determine the
turnover number (kcat) for β-galactosidase.

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Category: Sample Questions