Concepts and skills in the biochemistry/molecular biology lab

Authors


Abstract

Most colleges and universities throughout the world now offer a Biochemistry/Molecular Biology (BMB) lab course that is designed for undergraduate students in the molecular life sciences, chemistry, and related fields. To best serve our students, we must introduce them to the most current concepts, skills, and methods available. Suggestions for teaching a modern BMB lab are given here.

The Biochemistry/Molecular Biology (BMB)11 teaching laboratory has become a prominent and essential fixture in the training of undergraduate students for careers related to the molecular life sciences (biochemistry, molecular biology, chemistry, genetics, immunology, microbiology, neurochemistry, etc.). These students must acquire extensive experience working with biomolecules in the laboratory, and a formal lab course is usually the best first step to that experience. This step provides students the skills needed for future research participation at the undergraduate and graduate level and for jobs in the biotechnological and pharmaceutical industries. In addition, a lab experience is also an asset for those science majors preparing for careers in law and business that may be related but outside the realm of the basic sciences (patent law, pharmaceutical sales, etc.). With the acknowledged importance of a lab experience for all students, it is surprising that there is such a sparsity of literature references with discussion on the elements that make up an effective BMB laboratory experience. For example, what technical skills and procedures must be practiced and mastered by students? What teaching modes work best to most effectively train students in the lab? What instrumentation should undergraduate students become familiar with? What is the importance of the “other lab skills” such as communication (written and oral), teamwork, ethics, fairness, and responsibility? In this discussion I will not be able to provide completely satisfying answers to all these questions, but hopefully I will be able to provide insight and food for future thought.

BMB LABS OF THE PAST AND PRESENT

The standard approach to teaching the BMB lab, for many years, has been to:

  • select appropriate experiments from a textbook adopted for the class (Refs. 1–4 for current lab books), or

  • utilize a set of self-designed experiments, or

  • choose projects published in the biochemical education journals Biochemistry and Molecular Biology Education (BAMBED) and The Journal of Chemical Education (JCE).

These three options have provided instructors the opportunity to select lab activities that emphasize certain desirable biochemical principles, techniques, and skills and those methods that are compatible with the instructor's background/expertise and with the institution's instrumentation and facilities [5, 6].

However, major changes are now occurring in biochemistry and molecular biology, and these changes require that we adopt new instructional methods in the classroom and teaching lab [7, 8]. For example, the recent advent of bioinformatics, the merging of computer science with biology, is transforming the way we investigate protein structure and function as we can now obtain important information from databases and thus avoid days of “wet” work in the lab [9]. To facilitate the necessary changes in BMB education, Bell [7] suggests that we invoke the research paradigm that “Undergraduate courses should have investigative labs with realistic expectations about student involvement in experimental design, data analysis, and data interpretation.” While this approach may be desirable for those students who will later attend science graduate school, some instructors may prefer a more skill-based lab program for the average student. It is also important that labs have a multidisciplinary nature so students begin to experience the current merging of biology, chemistry, physics, math, and computer science.

The continuous development of new instrumentation and methods also will require changes in the way labs are presented. Now that protein characterization may be approached by NMR [10] and MS [11], should students struggle through an Edman degradation experiment in the lab just to see how it used to be done? Do we need to be concerned about the impact this will have on students who are at institutions that cannot afford the latest in instrumentation?

SELECT YOUR EXPERIMENTAL FORMAT

Many different teaching methods are now available for laboratory instruction, and professors may want to experiment with the new formats. Most lab courses in the past have been taught in the very traditional “skill-building” style. Sometimes called the “cookbook approach,” laboratory activities emphasize mastery of the primary tools and techniques of the trade and reinforce basic principles. Skill-building labs are characterized by the achievement of well defined goals and a very high success rate on the part of the students [12]. Students completing these labs often achieve high marks for technique but may show weakness in understanding the research process, the steps one goes through to solve a problem, and they miss out on the excitement of discovery. Most of the experiments published in lab manuals [14] and many in the education journals are in the skill-building mode [1315].

Discovery- (or inquiry)-based experiments display more active learning as they go beyond a “follow the recipe” approach. Students, who usually work in teams of up to four, are assigned a specific BMB problem to solve [16]. The student team is responsible for designing and completing experiments, integrating data, and finally arriving at reasonable conclusions that are reported in a written and/or verbal format. Students share the excitement of discovery (they experience “real” research), but they also must face the possibility of failure just as in research. Not all students in a lab course may be prepared for this independent approach so teams must be organized carefully.

Project-based experiments usually involve students working on a series of connected lab activities that revolve around a central BMB theme [17]. Students will have the opportunity to participate in all aspects of the project and learn many skills in the process. Although these are called “research-based” projects by some because of the continuity (current work is based on previous results), this term is somewhat misleading as students usually follow a preset schedule of activities and seldom waver from the plan. The central theme may be a biomolecule (e.g. enzymes and proteins are popular) or a biochemical process (e.g. recombinant DNA cloning techniques). Many of these projects originate in an instructor's research lab and make much sense at that instructor's institution. However, they often require unique and expensive resources and, since they are based on someone else's research, are logistically very difficult to transfer to other institutions. Instructors who set these up have extensive experience with the theme so they are very familiar with the system, its advantages, and its pitfalls.

Experiments that integrate all three of these teaching formats may be the wave of future [12, 18].

CONCEPTS AND SKILLS FOR THE FUTURE BMB LAB

No matter how sophisticated the instrumentation, how new the techniques, or how adept the instructor's pedagogy, a teaching lab without organized and thoughtful content is meaningless. We often turn to our scientific societies to gain advice on what principles and concepts students should learn.

The American Chemical Society divides recommendations for biochemistry lab into two categories [19].

Important general techniques:

  • error and statistical analysis of experimental data

  • spectroscopic methods

  • electrophoretic techniques

  • chromatographic separations

  • isolation and characterization of biological materials

Selected additional techniques:

  • use of radioisotopes

  • enzyme kinetics

  • immunoassay methods

  • DNA cloning and sequencing

  • plasmid isolation and mapping

  • peptide isolation and sequencing

  • computer graphics and structure calculations

The American Society for Biochemistry and Molecular Biology (ASBMB) has prepared a recommended curriculum for the undergraduate biochemistry degree, but it does not specify techniques or content [20]. The Education and Professional Development Committee of ASBMB is currently working on a new BMB curriculum that will include suggested lab skills.

The Biochemical Society (United Kingdom) has recently prepared a list of topic objectives to define the main content of the core curriculum for the Biochemistry First Degree [21]. Specific laboratory experiences mentioned in the report include:

  • analytical methods in chemistry (NMR, MS, etc.)

  • basic techniques for analyzing, cloning, and sequencing DNA

  • experimental techniques for the study and analysis of enzyme kinetics

  • techniques for studying macromolecular structure including purification and characterization and use of the computer for structural information

  • the main techniques used in cell biology

The Biosciences Industry Skill Standards Project (BISSP) sponsored by the United States Department of Education has recently generated an extensive list of skills expected of a bioscience technician [22].

The goal of all BMB lab instructors is to offer practical, hands-on experiences that introduce their students to the most contemporary instrumentation and principles the institution can afford. It is also important that students learn and practice all the steps necessary to design an appropriate experimental plan to solve a problem. Here I will attempt to present ideas on how these goals may be accomplished.

Table I shows “general” lab skills, procedures, and methods. This listing defines broadly those concepts that are used routinely and regularly in a lab setting for work on all types of biomolecules and for all types of measurements. This list is not all inclusive as instructors will be able to add a few of their own. The manner in which these general skills are presented by the instructor and practiced by the students will be dependent on many factors including the time and facilities available for the lab, the size of the class, the past experiences of the students, and the specific interests of the instructor. Perhaps it is helpful to provide some ideas on the mode of presentation and the relative amount of time spent on the general skills. In a typical BMB lab of 3–4 h per week, one-half of the first period may be spent discussing the first five skills listed in Table I (safety through computer). The remainder of the first period could be used for students practicing the next group of skills (solutions, pipetting, pH, buffers). Perhaps these skills could be learned with an experiment where students measure protein and/or nucleic acid solutions. I believe it is much more instructive to have students practice with “real samples” rather than just going through the motions of pipetting. Students at the BMB lab level will have already become adept at some of these general skills by work in earlier labs in Introductory Biology and Chemistry, Organic Chemistry, and others. Therefore it is important for instructors who know of the students' past experiences to make judgments about how much time should be devoted for each technique. It is expected that most students, after completing the BMB lab, would be proficient in the application of all of the general concepts in Table I. One exception to this may be the topic of radioisotopes where instructors need to make their own decision on the relative importance of this concept.

The importance of teaching skills in communication (writing and reporting results in Table I) is secondary only to the topic of lab safety. New scientific knowledge that is not communicated is of no value to anyone. Learning communication skills needs to begin with introductory labs (Chemistry and Biology) and continue through all future courses, lab and classroom [23]. Coordination and consistency among all lab instructors in a department are of vital importance so students do not learn different communication techniques at each level. Specific skills that students must master in the BMB lab include maintaining a lab notebook (journal), writing up a lab experiment, writing a journal-style article, critical analysis of other writing (other student's and journal articles), giving an oral presentation on experimental results, and the preparation and presentation of a poster [1, 12, 16, 18]. Students should practice presentations at their local institution and then gain experience at regional and national meetings.

Table II presents “selective” methods that serve a more specific purpose in the BMB lab as they may not be applicable for all types of biomolecules and measurements. It is obviously impossible for students to be introduced, in a one- or even two-term lab, to all of the selective lab methods. Instructors usually pick and choose those methods based on what facilities are available and what they believe their students should practice. The methods listed in Table II have been placed in order of relative importance on the bases of the author's prejudice and many years of experience. Methods that I would consider the most essential for BMB students include spectroscopy (UV/visible), chromatography (HPLC, affinity, gel filtration, ion-exchange, column), computational (enzyme kinetics/inhibition, ligand binding, databases), electrophoresis (all types), and biotechnology (recombinant DNA, plasmids, PCR, and restriction enzymes). Students should be encouraged to learn other skills and concepts in a future research experience.

Many of the methods listed in Table II require expensive instrumentation. Such equipment may not be present at smaller institutions, and even students at larger institutions may not have access to it because it is reserved for research. Instructors at these institutions must provide alternate opportunities to expose their students to the latest techniques and instrumentation. This may be done by visiting facilities at nearby research institutions, government labs, or industrial labs. Additionally, students could be encouraged to increase their understanding of the selective techniques by participating in a summer research project at an academic, industrial, or government laboratory. Students may also become acquainted with modern BMB principles and instrumentation by review of the University of Virginia Lab3D Project [24] and the Virtual Biochemistry Lab sponsored by the Nobel Foundation [25].

Science work is being done increasingly in groups, and students need to gain experience working in teams to gain confidence and to learn how to play the roles of team members. In addition to being adept with their hands, they also need to understand and practice the ethical characteristics of fairness, honesty, cooperation, and responsibility. This includes an understanding of fair and proper use of scientific data and the literature and a consideration of intellectual property, patents, and copyrights [26].

In these heady days when academic research and industrial science seem to be driven by instant results for the advancement of the principal investigator and for the profit motive, it is important that we and our students not lose site of the real goals of research and development: to better the lives of the peoples of the world.

Table Table I. General lab skills/procedures/principles
Lab safety
Writing and reporting results (maintaining a lab notebook, lab reports, posters, oral presentations)
Experimental design, collection and statistical analysis of data, controls
Reading the research literature with understanding
Computer (data analysis, graphing, spreadsheet, literature search, databases)
Preparation of solutions
Pipetting liquids
Buffers and pH
Measurement of protein and nucleic acid solutions
Isolation and/or characterization of biomolecules (amino acids, peptides, proteins, enzymes, carbohydrates, lipids, nucleic acids)
Centrifugation
Microfiltration/membranes and dialysis
Centrifugal vacuum concentration and lyophilization
Using commercial kits
Microarrays (nucleic acid and protein)
Radioisotopes
Table Table II. Selective lab methods
Spectroscopy: UV/visible, fluorescence, NMR (two-dimensional), MS (MALDI), FT-IR
MALDI, matrix-assisted laser desorption ionization; FT-IR, Fourier transform infrared; CE, capillary electrophoresis; IEF, isoelectric focusing.
Chromatography: HPLC, affinity, gel filtration, ion exchange, column, gas
Computational biochemistry: enzyme kinetics and inhibition, ligand binding, genomic and proteomic databases, molecular modeling, metabolic control analysis
Electrophoresis: PAGE (SDS and native), agarose, CE, IEF
Biotechnology: recombinant DNA, PCR, restriction enzymes, plasmid DNA, sterile techniques, growing bacteria, cloning, protein expression, gene libraries, sequencing DNA and proteins, blotting (Southern, Northern, Western, and other immunoassays)

Footnotes

  1. 1

    The abbreviations used are: BMB, Biochemistry/Molecular Biology; MS, mass spectrometry; ASBMB, American Society for Biochemistry and Molecular Biology; HPLC, high pressure liquid chromatography.

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