Exploring the Chamaecrista Gene Space

Student learning goals:

1. Frame a meaningful question that can be addressed using transcriptome data.
2. Propose a testable hypothesis based on reading several journal articles that can be tested using transcriptome data.
3. Modify a hypothesis based on data analysis.
4. Use multiple bioinformatics resources without losing focus on a biological question.
5. Distinguish types of information that can be extracted from a transcriptome sequence vs. a genome sequence.
6. Demonstrate ability to leverage investigation by using the whole transcriptome for analysis (i.e. pattern search).
7. Understand that SNPs are mutations.
8. Use mutant analysis in a related species (e.g. garden pea or Arabidopsis) to formulate hypotheses based on SNP variation in ecotypes.
9. Design an experiment that can be tested using PCR based on a working hypothesis derived from transcriptome analysis.

The Chamecrista activity takes place over 8-9 weeks of laboratory in an undergraduate genetics course.The first two weeks are focused on working with whole plants and framing questions and hypotheses that can be addressed with in silico data during the following two weks. The in silico analysis is used to develop questions and hypotheses that can be experimentally tested during the next three weeks. Students then give presentations and use feedback to prepare a final group paper on all three aspects of their work. It would be possible to modify any of the modules as stand alone activities.

Students participating in this 4 h per week lab course must also be enrolled in the genetics course which is taught by the same instructor. Key concepts in lab are integrated into the flow of instruction in the course. The prerequisite for the course is a two term introductory biology course with associated labs.

Students are encouraged to explore genes associated with the interactions between environment and flowering because of the instructors expertise. The focus could readily be modified to match the interests of other instructors.

Students are encouraged to choose among several strategies for the in silico lab. An alternative would be to chose a single strategy for all students (e.g. a candidate gene approach).

Whole Plant Investigation

The first two weeks of the lab connect the students with whole plants and large phenology data sets. Phenology is the study of the timing of naturally occurring events such as the date when the first leaf of a plant expands or the first flower opens (first bloom).
Lab handout for students for the whole plant experiments (Microsoft Word 29kB Jun22 09)

In Silico Investigation

During weeks 3 and 4, students use the Chamaecrista Genome Explorer to ask questions that can be answered using sequence, expression, and variant data for Chamaecrista transcriptomes. Students can complete this part of the activity without having worked with the whole plants. Only minor modifications to the Lab handout for students for the in silico investigation (Microsoft Word 45kB Jun22 09) are needed. 

PCR-based Investigation

Students will need to design primers to address their question, isolate RNA, quantify their RNA, use one-step RT PCR to make cDNA and amplify the cDNA, run a gel, and capture an image of their gel. The following handouts may be useful to your students, although they will need to be adapted for the equipment you have in your labs. During week 5 student isolate their RNA. Week 6 is used to quantify the RNA using a spec and setting up the PCR reaction. We use week 7 to separate the PCR products on a gel and to capture the images and do an analysis of the relative intensities of the bands.

Tips for Primer Design (Microsoft Word 2007 (.docx) 137kB Jun19 09)
RNA Isolation and Determination of Concentration (Microsoft Word 36kB Jun19 09) - We use a FastRNA Pro Green Kit from MP Biomedicals which requires a Thermo FastPrep 24 Homogenizer. 
RT PCR (Microsoft Word 81kB Jun19 09)
Visualizing Your PCR Products with Gel Electropheresis (Microsoft Word 2007 (.docx) 109kB Jun19 09)

Growing Chamaecrista plants from seed (Microsoft Word 35kB Jun19 09)
Setting up the lab for RNA isolation (Microsoft Word 57kB Jun22 09)
Setting up the lab for PCR (Microsoft Word 56kB Jun22 09)
Tips for the RT PCR lab (Microsoft Word 2007 (.docx) 133kB Jun19 09)

We used the following assessment tools:

ACT CAAP Science Reasoning (pre) - http://www.act.org/content/act/en/products-and-services/act-collegiate-assessment-of-academic-proficiency/test-prep.html#ScienceTest

ACT Science Reasoning

Genetics Concepts Assessment (pre/post) - Smith, M.K., Wood, W.B., and Knight, J.K.(2008) The genetics concept assessment: A new concept inventory for gauging student understanding of genetics. CBE - Life Science Education. 7: 422–430.

Motivated Strategies for Learning Questionnaire(MSLQ, pre) - 
Pintrich, P. R., Smith, D.A., Garcia, T., & McKeachie, W. J. (1991). A 
manual for the use of the motivated strategies learning questionnaire 
(MSLQ). (Tech. Rep. No. 91-B-004). Ann Arbor: University of Michigan, 
National Center for Research to Improve Postsecondary Teaching and Learning

Washington State Critical Thinking Rubric (applied to two papers, powerpoint presentation) - libarts.wsu.edu/genstudies/undergraduate/ctguide.asp
Adaptation of Critical Thinking Rubric for Chamaecrista presentation and papers (Microsoft Word 2007 (.docx) 112kB Jun19 09)

Journal entries

Web click throughs

Classroom observers from SERC

If you choose to use the whole plant component of this activity, you can obtain seeds for the 26 Chamaecrista fasciculata accessions from the USDA. If you would like some of the MN, KS, and OK seeds, please contact Susan Singer (ssinger@carleton.edu).

For background information, journal articles are listed in the student handouts for both the whole plant investigation and the in silico investigation. The articles by Julie Etterson will provide a broader context for questions about the MN, KS, and OK ecotypes.