GENETIC ANALYSIS AND BIOTECHNOLOGY - BIOLOGY 110

4.00 units (CSU transferable; UC transferable) Summer 2018 Professor: Patricia Zuk, PhD email: [email protected]

LECTURE: Section 11862 MSA Rm. 303 Monday, Tuesday, Wednesday, Thursday 11:30 AM to 1:35 PM

LABORATORY: Section 11868 MSA Rm. 303 Monday, Tuesday, Wednesday, Thursday 1:35 PM to 3:40 PM

OFFICE HOURS: by appointment or from: 3:45 to 4:45 PM Monday through Thursday Office is MSB Room 210

PREREQUISITES: Biology 6, Chemistry 101 and Math 125 with a grade of ‘C’ or better in each course. High school AP biology and chemistry or introductory college biology and chemistry may also be accepted with permission of instructor. However, it is strongly recommended that you take Biology 6 before taking this class.

COURSE DESCRIPTION: Biology 110 is designed for Biology Majors wishing to transfer to the UC and CSU systems and pursue a degree in the life sciences. This course provides a comprehensive introduction to genetic analysis whereby students examine topics such as genomics, chromosome analysis and population genetics. This course also provides a comprehensive introduction to the science of biotechnology by providing both the theory and hands-on experience with current laboratory procedures.

ATTENDANCE: Attendance is mandatory (see Administration Regulation E13). If enough absences occur throughout the semester, I can exclude you from the course. Be aware that your grade in this course depends on your performance – which is dependent upon your attendance. I guarantee if you miss too many classes and labs – you will NOT do well in this course.

Since biology labs cannot be duplicated outside the class it is very important for you not to miss any labs if possible. You also must plan on attending the entire lab period. When you are finished the labs – to my satisfaction – you may leave quietly without disturbing your fellow lab mates.

I consider extreme tardiness or early departure from lab/lecture without a valid cause to be very disrespectful conduct. However, I realize traffic and life gets in the way sometimes. So being late and having to leave early is fine – every now and then. Do NOT insult me or your classmates by consistently showing up late to lecture/lab every time!!!

DO NOT EVEN CONSIDER BEING LATE IF THERE IS AN EXAM SCHEDULED. I will NOT give you the exam if you are more than 10 minutes late and have provided me with a valid excuse for your tardiness that day!! If you have conflicts in your schedule – come and talk to me. I am very understanding about many things and do not bite my students (much!). Also, exchange numbers with your lab-mate so that if you are running late for an exam you can relay a message to me through them.

WITHDRAWING FROM THE CLASS: Any student withdrawing from the class must inform the admissions office and complete the required steps. Students failing to follow the correct procedure for withdrawing will receive an ‘F’ at the end of the semester. I will not be held responsible for your grade if you fail to correctly withdraw from this course. Therefore, confirm your registration status. Finally, there are deadlines for withdrawing without a “W”, with a “W” and a deadline where withdrawing is no longer possible. Be aware of these dates.

COURSE CONSTRUCTION: This course is comprised of four weekly lectures/labs that total over 16 hours per week! This is a lot of lecture time and a lot of lab time. Therefore, several short breaks will be given throughout the evening. But be aware that when running a lab, a break may not be possible.

You are welcome to tape my lectures. I also have my own personal website – www. patriciazuk.com where the lecture presentations can be found along with additional learning materials. This website is password protected with the username of student and the case-sensitive password of #1Wlacstudent. Be sure to use a capital “W” or you will not be able to access the website. The lectures on this site are “student lectures” and do NOT contain every detail you will find in my lecture presentations or will hear throughout my lectures. This is so that you are required to pay attention and write some things down. Therefore, please print out these lectures and bring them to class so that you may supplement them throughout the lecture/lab period with your own notes taken during class. You will also be required to re-create simple figures and diagrams that I will present to you throughout lecture.

Videos shown in lecture and lab are to be considered as important as lecture and you should pay close attention to the material presented in them.

Handouts will be given in class so be sure to pick them up the day they are offered. I am not guaranteeing that these handouts will be available after the day I offer them.

LABORATORIES: Several labs that demonstrate important concepts will be given in this course. These labs are detailed in the Biology 110 Laboratory Manual that is on sale at the bookstore. Each student MUST buy the course lab manual. Make sure to bring your lab manual and all lab handouts to each lab as your assigned material will be in these materials. If no lab is planned, then another lecture will be given during that time.

WEST LA COLLEGE STUDENT LEARNING OUTCOMES (SLOs): West LA College as an institution is committed to an environment of learning and respect for its students. Its mission is to serve the community by providing quality instructional services through its programs and facilities. The college has created a series of Student Learning Outcomes (SLOs) that are designed to maximize the successes and experiences of the students here at WLAC. A. Critical Thinking: Analyze problems by differentiating facts from opinions, using evidence, and using sound reasoning to specify multiple solutions and their consequences. B. Communication: Effectively communicate thought in a clear, well-organized manner to persuade, inform, and convey ideas in academic, work, family, and community settings. C. Quantitative Reasoning: identify, analyze, and solve problems that are quantitative in nature. F. Technological Competence: Utilize the appropriate technology effectively for informational, academic, personal, and professional needs.

BIOLOGY PROGRAM SLOs: In addition, the Biology program also has several unique SLOs. A student who completes this program will be able to: 1. Explain how scientists investigate causes of natural biological phenomena. 2. Explain how living things are organized, reproduce, acquire matter & energy, and inherit & express genetic instructions. 3. Utilize biological information to make informed decisions about environmental issues. 4. Utilize biological information to make informed decisions about personal issues. 5. Perform basic biological lab procedures.

STUDENT LEARNING OUTCOMES FOR BIOLOGY 110: At the end of the semester, the students should understand and be able to explain the fundamental concepts of the following:

1. What a genome is 2. Mendelian genetics and the chromosomal basis of inheritance including the laws of segregation and independent assortment 3. What is a genetic mutation and where do they occur?

SUBJECT OBJECTIVES: At the end of the semester the students should demonstrate proficiency in understanding and explaining the following: 1. The structure and function of the nucleus, including how DNA is organized in both prokaryotes and eukaryotes. 2. The process of DNA replication and RNA transcription and protein translation in prokaryotes. 3. The control of prokaryotic gene expression. 4. The structure of a bacterial plasmid and how it can be used in recombinant DNA technology 5. How bacteria can be transformed with recombinant plasmids 6. The principles of amplifying DNA through polymerase chain reaction (PCR). 7. How restriction enzymes work, including what sequences of DNA do these enzymes recognize and how they can be used in the creation of a piece of recombinant DNA. 8. The types of ligations that can be used in creating recombinant DNA. 9. How agarose gel electrophoresis separates DNA pieces of different sizes.

TECHNICAL OBJECTIVES: Add the end of the semester, the student should be able to perform the following within a laboratory setting: 1. Weighing a given substance using an electronic balance. 2. Determining the absorbance of DNA using a spectrophotometer and the ability to calculate its concentration based on this value. 3. The ability to pipette specific volumes using serological pipets and a pipet-aid or a micropipette 4. The set up and performance of an experiment to isolate of DNA from bacterial cells 5. The set up and performance of an experiment to analyse isolated DNA, including being able to make an agarose gel, run the DNA using that gel and analyze the resulting DNA migration pattern 6. The set up and performance of an experiment to analyse isolated DNA by means of a restriction digest. 7. The set up and performance of an experiment to amplify DNA by PCR. 8. The set up and performance of an experiment to purify genomic DNA 9. The simulation of DNA replication, RNA transcription and protein translation if given specific DNA sequences.

COURSE MATERIALS: be sure to bring these to each class

1. Textbook: There is no required textbook. Any used genetics textbook will be fine. Some suggestions are given below a. Genetics: A conceptual approach. Benjamin A. Pierce. b. Genetics: From Genes to Genomes. Leeland Hartwell & Michael Goldberg. c. Genetics: Analysis & Principles. Robert J. Brooker

2. Lab Manual: Available in the bookstore. You MUST purchase this lab manual as it contains all of the labs you will perform in this course.

3. Lecture notebook: This book will be used to supplement the lectures given in the morning and afternoon sessions. You may also want to print out the lecture slides prior to coming to class and put these in your lecture notebook. As a result, a three-ring binder may be a good option. That way you can place your notes and the printed slides together in the same notebook along with any handouts given during lecture.

4. Numerous colored pens and pencils for lectures and labs

5. Scantron 882E forms for exams

EXAMINATIONS: You will have 3 lecture exams each worth 100 points for a total of 300 possible points. All lecture exams will be based on my lectures and will also test you on the techniques and the concepts you have learned in your labs. This means that the materials in the lab manual will be on the exam. Types of questions may include multiple choice, fill in the blank, short answers and genetic word problems. Lecture exams will range anywhere from 50 to 100 questions but will be converted to 100 points.

I will discuss each exam and what to expect– so don’t freak out! I may also provide you with some study guides to ensure you are keeping yourself on track during your study times. But don’t count on it! This is a majors level biology course so you are expected to know what could be on an exam.

I do not allow you to keep any of your tests so please keep track of your performance in the class by recording all your exam scores.

Cheating will NOT be tolerated. ANY STUDENT FOUND CHEATING WILL RECEIVE THE GRADE OF ‘F’ FOR THAT EXAM AND MAY BE EXPELLED FROM THE COURSE!!! Please see the college’s policy on academic dishonesty for additional information. While not written in this syllabus, the college’s policy on academic dishonesty will be adhered to in this course. Schedule of Topics Text Week Date Lecture Topic Chapters Lecture #1: An introduction to Genetics 06/11 Pipetting Lab Lecture #2: How Genes Work: DNA replication 06/12 Genomic DNA Isolation Lab Lecture #2: How Genes Work continued: RNA 1 transcription and Protein translation 06/13 HOMEWORK: Lecture #3: Review of Mitosis, Meiosis and the Cell Cycle Lecture #4: Anatomy and Function of a Gene: Dissection 06/14 through Mutation Agarose Gel Electrophoresis Lab Lecture #5: Anatomy and Function of a Gene: 06/18 Transposable elements, DNA mutagens and DNA repair Replica Plating Lab Lecture #6: How Genes are Regulated: Gene expression 06/19 in prokaryotes – the operon model of gene expression 2 DNA Electrophoresis Lab preparation Lecture #7: How Genes are Regulated: Control of 06/20 eukaryotic gene expression DNA Electrophoresis Lab & analysis Lecture #7: How Genes are Regulated continued: 06/21 Control of eukaryotic gene expression EXAM 1 06/25 12:00 PM to 3:00 PM Lecture #8: The Eukaryotic Chromosome: Euchromatin & Heterochromatin, DNA condensing, Transcriptional 06/26 silencing 3 Restriction enzyme Lab preparation Lecture #9: Mendelian Genetics: The two laws and the 06/27 Chromosomal Theory of Inheritance Restriction enzyme Lab Lecture #10: Extension of Mendelian Genetics: 06/28 Dominance, Pedigrees & Gene interactions Restriction enzyme Lab analysis Lecture #11: Linkage & Mapping: Linkage & Chi square 07/02 analysis Lecture #12: Genetic variation: Crossing over and 07/03 Recombination frequency 4 07/04 NO CLASS

07/05 Lecture #13: Genetic variation: Mapping Linked Genes EXAM 2 07/09 12:00 PM to 3:00 PM Lecture #14: Genetic variation: Complementation assays 07/10 in bacteria, the rII locus of bacteriophage 5 Lecture #15: Genetic variation: SNPs and RFLPs 07/11 PCR Lab Lecture #15: Genetic variation continued: SNPs and 07/12 RFLP PCR Lab analysis Lecture #16: Bacterial Genetics: conjugation, 07/16 transduction, DNA cloning & complementation assays Bacterial Transformation Lab Lecture #17: Organelle Genetics & Non-Mendelian 07/17 Inheritance 6 Bacterial Transformation Lab analysis Lecture #18: Human Genetics: genetics & evolution, 07/18 population genetics, the Hardy-Weinberg equation EXAM 3 07/19 12:00 PM to 3:00 PM

Lecture Topic #2 – How Genes work – DNA replication, Transcription and Translation Nucleic acids review DNA structure Nucleus structure – nucleoplasm, nucleolus & chromatin Chromatin organization and condensing DNA replication mechanism -DNA polymerase III structure and function -helicases and topoisomerases – role in replication -primase function in replication -replication of the sense vs. anti-sense strand – Okazaki fragments One gene, one enzyme hypothesis and experiments in Neurospora crassa RNA structure vs. DNA Types of RNA Transcription mechanism: RNA polymerase II – structure and function mRNA modifications Splicing mechanism, snRPs and the spliceosome Alternative splicing – advantages Codon composition Intragenic suppression in the determination of the codon Third codon wobble tRNA and ribosome structure and function in translation Charging a tRNA molecule Formation of the prokaryotic and eukaryotic translation initiation complex -RBS, Shine Delgaro and Kozak sequences -reading frames Mechanism of translation

Lecture Topic #3 – Mitosis, Meiosis and the Cell Cycle Differences between Mitosis and Meiosis Spermatogenesis vs. oogenesis Phases of the Cell cycle – events and function Phases of Prophase I: zygonema, pachynema, diplotene, diakinesis Synapsis & Crossing over Crossing over patterns – two, three and four-strand

Lecture Topic #4 – Anatomy and function of a Gene: Dissection through mutation Allele frequency – definition and calculation Mutation types: definition and examples -transversion -transition -inversion -point mutation types -insertions and deletions -chromosomal rearrangements -translocations -loss and gain of function mutations Mutation promotion of natural selection

Lecture #5 - Transposable elements Retroposons and transposons Mechanisms of retroposon and transposon function Spontaneous mutations and bacteria and bacterial resistance Types of mutagens Chemical and physical changes to the DNA DNA repair mechanisms -mismatch repair, excision repair -repair mechanisms in cancer -recombination mechanisms Tri-nucleotide repeats and fragile X disease – impact of repeats on disease development Complementation testing to determine presence of mutations

Lecture Topic #6 – Gene regulation in prokaryotes - Prokaryotic gene expression Operon definition and types Inducible vs. repressible operons Lac operon – structure & function -Gal and permease function in lactose metabolism Inducers and repressors – definitions and roles in the operon Activators and repressors as DNA-binding proteins – domains of a DNA-binding protein Operator region and rotational symmetry Allosteric regulation of inducers and repressors Co-repressors – definition, functions and examples Pajamo experiment lacIs, lacI- and lacOc mutations – functions and effect on the operon trans vs. cis acting factors – experimental evidence (i.e. the cis-trans test) IPTG and X-Gal use in labs – blue-white screening Positive gene regulation of the lac operon – CAP/CRP protein and cAMP Dual control of the lac operon – glucose vs. lactose Ara operon – function and structure (CAP site, I site) Positive and negative regulators of the ara operon

Lecture Topic #7 – Gene regulation in eukaryotes – Eukaryotic gene expression Why is eukaryotic gene expression more complicated? Stages of eukaryotic gene expression regulation Organization of a typical eukaryotic gene Enhancers and promoters – function in expression Transcription factors – general vs. activators vs. repressors -structure and function in regulation of gene expression Response elements and activators in gene expression Use of reporter genes in promoter function Eukaryotic promoter construction – the core promoter – TATA boxes and CpG islands -promoter proximal elements (PPEs) – Oct sequence, GC-rich regions & the CCAAT box Enhancer construction – location in the gene -sequence elements -activator binding The transcription Initiation Complex (TIC) – composition and function in transcription The pre-initiation complex – formation and role in transcription DNA bending protein in creation of the transcriptional initiation complex Three ways activators increase gene expression Homo vs. heterodimers DNA binding motifs of activators – helix-loop-helix, helix-turn-helix, leucine zipper, zinc finger Function of co-activators in the TIC Repression mechanisms – competition vs. quenching Role of chromatin organization in transcriptional silencing Post-transcription control -mRNA processing -cap and tail function -splicing mechanism, snRPs and the formation of the spliceosome -5’ and 3’ UTR structure and function -3’ UTR and poly-adenylation Alternative splicing – role in Drosophila sex development Translation Initiation – formation of the Initiation Complex (ribosome binding) Post-translational processing – modifications to the polypeptide by chemical groups Non-coding RNA – miRNA & siRNA – control over gene expression

Lecture Topic #8: The eukaryotic chromosome Euchromatin vs. heterochromatin Histones and nucleosomes – chemical modifications DNA condensing mechanism – the radial loop model Mechanisms of transcriptional silencing – position effect varigation X-inactivation – mosaicism vs. chimeras Satellite DNA Centromeres – types, locations and function Telomeres

Lecture Topic #9: Mendelian Genetics Mendel’s experiments – pea plants, P and F generations Law of Segregation Mendelian model of inheritance Genotypes, phenotypes, alleles Punnett squares and test crosses Pedigree analysis Law of Independent Assortment – monohybrid vs. dihybrid crosses Laws of probability – monohybrid crosses -multiplication rule – i.e. the Product rule -addition rule – i.e. the Sum rule Probability and dihybrid crosses

Lecture Topic #10: Extensions of Mendelian Laws – Inheritance patterns and their complexity What are the three degrees of dominance? How do incomplete and complete dominance come about biologically? What is a polymorphic gene? How can Tay-Sachs illustrate degrees of dominance? Pleiotropy Multi-allelelic traits and the dominance series in rabbit coat color and lentils Multi-factor traits – how do they differ from Mendel’s findings? Genotypic classes and the determination of lentil seed color Epistasis and Complementation -what is the biochemistry underlying these phenomena? -what phenotypic ratios do they produce? -recessive vs. dominant epistasis Penetrance vs. expressivity The action of modifier genes Conditional lethal mutants – permissive vs. non-permissive conditions Phenocopy Polygenic inheritance – continuous and discontinuous traits Sickle cell anemia as an example of the extension of Mendelian laws

Lecture Topic #11– Linkage and Chi-square analysis Color-blindness and Hemophilia A and B – linked vs. non-linked genes on a chromosome Three themes of linkage How do linked genes assort? Linkage of autosomal traits vs. sex-linked Confirmation of linkage – chi-square analysis Goodness of fit Other uses for chi square analysis

Lecture Topic #12 – Crossing over and Recombination Frequency Recombination – definition Calculating Recombination frequency Sources of recombination Experimental evidence for crossing over – fruit fly studies Cohesins and Anaphase I vs. Anaphase II – chromosomal segregation Chiasma and breakpoints – definition Mechanism of crossing over -the heteroduplex region -the double strand break model -resolution of the Holliday junction – cross over and non-cross over products Genetic maps and calculating map distances on a chromosome – using recombination frequencies to map genes on a chromosome -examples from Thomas Hunt Morgan and Alfred Sturtevant’s studies -using two and three point crosses Parental vs. Recombinant classes and the calculation of map distances Calculating the coefficient of coincidence Calculating interference due to double cross-overs Mitotic recombination and genetic mosaics

Lecture Topic #13 – Crossing over and Recombination Frequency Mapping – definition Chromosomal mapping & recombination frequency Using recombination frequency to determine map distances – 2 and 3 point crosses

Lecture Topic #14: Complementation assays in bacteria Intragenic recombination Seymour Benzer – complementation testing of the rII locus in bacteriophage -T4 life cycle – plaque formation -infection of E.coli strains -experimental protocols using T4 Benzer’s T4 complementation assays -outline of approach -differential results in E.coli strains -confirmation of T4 infection Complementation groups – definition -interpretation of complementation assays - determining the locations of mutations on different genes vs. the same gene Determining complementation groups from complementation tables Deletion mapping – how does it work? Fine mapping of the rII locus

Lecture Topics #15– DNA polymorphisms What is a DNA polymorphism and a polymorphic locus? 5 major types of DNA polymorphisms Use of SNPs in positional cloning of a disease gene How are SNPs detected? - RFLP analysis and microarrays -what is a Restriction enzyme? Restriction site? DIPs – definition Simple sequence repeats (SSRs) – definition -detection through PCR -PCR amplification of DNA – the Taq polymerase, primer design, how does PCR mimic DNA replication? -SSRs and DNA fingerprinting -CODIS and forensic use of STR/SSRs What is positional cloning? The use of DNA polymorphisms in positional cloning

Lecture Topic #16 – Bacterial Genetics Phylogeny of bacteria – major groups E. coli characterization -Growth conditions and growth curves -K12 and B strains Genome organization – gene categories Plasmids Mutational studies in bacteria – types of mutations Genetic screens Blue-white screening Gene transfer in bacteria – 3 types Transformation – natural vs. artificial -Competent bacteria and electroporation Conjugation – F plasmids and HfR strains Transduction – generalized vs. specialized -Bacteriophage – recombinant and prophage -Lytic vs. Lysogenic cycles -Transduction and mapping

Lecture topic #17 – Organelle Genomes and Non-Mendelian Inheritance Chloroplast vs. mitochondria structure Endosymbiotic theory of evolution – ancestors of mt and cpDNA Heteroplasmy vs. homoplasmy Mitochondrial genomes -Size and organization – human vs. yeast -Nucleiods -Replication mechanism -Kinetoplast organization -Mini vs. Maxicircles RNA editing of mtDNA transcripts Mitochondrial exceptions to the genetic code Non-Mendelian inheritance of mtgenome -Uniparental inheritance Mitochondrial mutations and diseases -LHON -MERFF

Lecture Topic #18 – The Human Genome & Population Genetics Definition of a genome How is a genome organized? Bacterial vs. animal vs. human vs. organelle The human genome project – approaches for mapping a genome -shotgun vs. whole genome approach Heterozygote superiority 4 agents of evolutionary changes in populations – mutation, migration, genetic drift & natural selection Recessive allele frequency in populations - genetics and evolution Hardy-Weinberg equilibrium

LAB TOPICS TO BE COVERED

Analysis of DNA i. restriction enzymes – natural role and use in labs; sticky vs. blunt ends ii. inactivation of restriction enzymes iii. blunt vs. sticky ends iv. gene sequencing – dedeoxy method vs. automated sequencing

Gene cloning & the study of gene expression – how do you affect the level of protein expression? i. plasmid construction ii. ligations iii. miRNA regulation of gene expression iv. gene knockdown – siRNA knockdown v. cloning and recombinant plasmids vi. cloning, GMOs and transgenic animals – benefits and pitfalls

Forensics and biotechnology i. PCR amplification of DNA – primers and oligonucleotides ii. polymerase creation of synthetic DNA – vs. natural polymerases iii. DNA fingerprinting – RFLPs and SSRs