Download Activity 1 - Assignment 2 - Workforce Development in Stem Cell

Module 2: From Embryo to Beating Heart
Section 4: Directed Differentiation of hESCs into Endoderm
2.4.2: In class group activity
Assignment 2: Group Activity - Planning an experimental approach to
test the efficiency of directed differentiation of human ESCs into
endoderm.
Goals:
At the end of this exercise, students will be able to:
1) Work as a group to identify a question and plan an experimental approach to address
that question.
2) Identify and discuss about the advantages and disadvantages of potential experimental
methods.
3) Determine which experimental approach will be the most appropriate to address the
specific question.
4) Plan and prepare a written protocol for the experimental procedure to be used in the
laboratory for this purpose.
Rationale:
The process of scientific research involves multiple steps that range from: a) identifying
an important question, b) planning the experiments that would address the questions, c)
gathering and sorting out the data, d) synthesizing and evaluating the information and e)
reporting about the scientific findings.
Understanding how to approach each of these steps is essential not only for the success of
the experiments but also for building a logical framework about everyday work in a
research laboratory. Teamwork is an essential component of the scientific research since
it engages us in discussions that may help to solve scientific problems. Various members
of the team bring different perspectives on either how to solve a problem, or what
experimental procedures to use to answer a specific question, that you may have not
thought about on your own. Therefore, a group discussion about planning your
experimental procedures is vary valuable before you embark on the wrong procedure that
will not give you an answer and waste your time, resources and energy.
Instructions:
1) You should bring in the class your completed individual worksheet from Assignment
1 since this will serve as a guideline for group discussion and for planning your
experimental procedure in the laboratory.
2) The instructor will arrange each student in groups of three before the beginning of
this activity. The instructor will assign one student to present the opinion of the group
in front of the class. The other members of the group will take notes and prepare to
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Module 2: From Embryo to Beating Heart
Section 4: Directed Differentiation of hESCs into Endoderm
2.4.2: In class group activity
answer questions from the instructor or other groups about their proposed
experimental procedure or why they think that this is the most useful method to
address the question.
3) You will discuss for 30 minutes and fill in a group worksheet that has identical
questions to the individual worksheet from the Assignment 1 about experimental
procedures that you are planning to use to determine the efficiency of the directed
differentiation protocol. However this group worksheet should represent the
consensus for the group and it should address the following questions:
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Which markers define each cell population (e.g. ES cells, EBs, immature
endoderm or definitive endoderm) and when do these markers appear during
differentiation?
How can you use this information to design an experiment?
What is the best experimental procedure to analyze the outcome of this
experiment? You should prepare a list of advantages and disadvantages for your
experimental approach to help you answer this question.
What types of controls are you going to use for your experiments?
What is the proportion of various cell types that you should expect at various days
of the directed differentiation procedure?
How are you going to determine the proportion of each cell type in your
experiment?
4) At the end of the discussion, the selected representative from your group will present
the group’s summary in front of the class for 5 minutes.
5) The other members of the group should be prepared to answer any questions from the
instructor or other groups that will be asked at the end of the presentation (5 minutes).
6) Give your instructor the individual and group worksheets for evaluation.
7) The instructor will decide at the end of all presentation what is the best experimental
approach to use in the laboratory for analyzing the efficiency of directed differentiation
of human ESCs into endoderm.
Reading Materials:
Please refer to the reading material list from Assignment 1 in preparation for this
assignment.
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Module 2: From Embryo to Beating Heart
Section 4: Directed Differentiation of hESCs into Endoderm
2.4.2: In class group activity
Worksheet: Planning an experimental approach to answer a question.
Group #
Student 1:____________________________
Student 2:____________________________
Student 3:____________________________
Step 1: The following diagram represents the differentiation of ES cells into endoderm
in culture. Think about the following small questions and fill in Table 1 (Please review
the reading notes to answer the following questions and fill in Table 1).
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What markers characterize each cell type in this diagram?
When are these markers expressed?
When do these cell types appear in the dish during the directed differentiation
protocol?
What do you expect the frequency for each cell type to be at the end of your
protocol?
TABLE 1
Cell type
ES cell
Embryonic
Body (EB)
Molecular
Markers
Time (days) of
appearance in
the dish during
3
Immature
endoderm
Definitive
endoderm
Module 2: From Embryo to Beating Heart
Section 4: Directed Differentiation of hESCs into Endoderm
2.4.2: In class group activity
differentiation
Frequency
at
the end of the
differentiation
Step 2: You should think how to use the information that you gathered in Table 1 to plan
your experiment.
a) What possible experimental procedures can you use to address the question?
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b) What are some advantages and disadvantages for each of the procedures?
Experimental procedure # 1 (name) :__________________________________________
Advantages
Disadvantages
1.
2.
3.
4.
5.
Experimental procedure # 2 (name) :__________________________________________
Advantages
Disadvantages
1.
2.
3.
4.
5.
c) Briefly describe which method would be most appropriate for the experiment and
why?
________________________________________________________________________
________________________________________________________________________
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Module 2: From Embryo to Beating Heart
Section 4: Directed Differentiation of hESCs into Endoderm
2.4.2: In class group activity
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d) What positive and negative controls can you use for your experiment?
Positive control
Negative control
Additional controls
e) What concentration are you going to use for the reagents?
________________________________________________________________________
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f) How much time do you need to complete the experiment and why?
________________________________________________________________________
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g) How are you going to gather your data?
________________________________________________________________________
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h) How are you going to analyze and organize your data?
________________________________________________________________________
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Module 2: From Embryo to Beating Heart
Section 4: Directed Differentiation of hESCs into Endoderm
2.4.2: In class group activity
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