Course Overview: Students will learn and apply molecular biology techniques (focused on nucleic acids) in the laboratory. Students will learn principles and practice of basic bacterial culture techniques, transformation, agarose gel electrophoresis, nucleic acid purification (plasmid and genomic DNA, RNA), nucleic acid quantification, DNA restriction digestion and analysis, polymerase chain reaction (PCR), and basics of computer-based DNA sequence analysis and planning of cloning strategies. In addition, students will learn about the nature and selection of DNA cloning vectors (bacterial & eukaryotic), restriction enzymes, modifying enzymes, polymerases, and other reagents used in molecular biology. We will examine aspects of bioinformatics and genomics, and newer/advanced molecular technologies such as next-generation sequencing and microarrays that are especially important in genomics, and discuss the burgeoning other '-omics' fields. Some primary literature articles illustrating standard techniques will be studied. We will apply our newly learned molecular techniques toward solving real biological research questions, and to carry out a basic DNA cloning project.

Syllabus – Deadlines - The timing in this document are guidelines.  Some experiments may take more time, others less.  Due dates

for assignments will be provided in class.  The web will have target dates for all assignments as well as a description for each

homework assignment.

	Tuesday Lab 8:00 – 12:00 (Lab)		Thursday Lecture/Lab 10:45 – 12:05 (ST 130)
		Sept 7	Block 1 Lect: Intro, lab safety, electronic notebooks, solutions, Lab math, intro to sterile and bacteriological tech
Sept 12	Block 1 Lab 1: Pipetting and Culture Tech.  Discussion: Pipetting, bact culturing / media prep and autoclave training	Sept 14	Block 1 Lect: bacterial strains, antibiotics and transformation, Plasmid purification,
Sept 19	Block 1 Lab 2: Transformation Discussion: Bacterial Plasmids Promoters, DNA electrophoresis, DNA/RNA quantification	Sept 21	Block 1 Lect: SOP/QA/QC, working in industry
Sept 26	Block 1 Lab 3: Plasmid Purification & quantification	Sept 28	Block 1 Lect: Restriction digest, intro to cloning and subcloning.
Oct 3	Block 1 Lab 4: Agarose Gel & Restriction Digest	Oct 5	Block 1 Lect: In class project.  Cloning Assignment I and II.
Oct 10	Block 2 Lab 1: Bioinformatics and sequence analysis workshop	Oct 12	Block 2 Lect: Next-generation and advanced high throughput sequencing.
Oct 17	Block 2 Lab 2: Trip to biotech/pharma company	Oct 19	Block 2 Lect: Catchup and Data Analysis
Oct 24	Block 2 Lab:SNP analysis of human tissues	Oct 26	Block 2 Lect: SNP analysis of human tissues
Oct 31	Block 2 Lab:SNP analysis of human tissues Block 2 Lab 3: Advanced Cloning Techniques - Advanced PCR theory and techniques, primer design, site directed mutagenesis, sequence insertion and qPCR	Nov 2	Block 2 Lab 3: Continued - Advanced PCR theory and techniques, primer design, site directed mutagenesis, sequence insertion and qPCR
Nov 7	Block 2 Lab: rtPCR expression of NHE1	Nov 9	Block 2 Lab: rtPCR expression of NHE1
Nov 14	Block 2 Lab: rtPCR expression of NHE1AND Block 3: Cloning Project Design/Introduction	Nov 16	Block 3: Cloning Project Design/Introduction
Nov 21	Block 3: Cloning Project Planing and Start Project	Nov 23	Thanksgiving
Nov 28	Block 4: Cloning Project	Nov 30	Block 4: Cloning Project
Dec 5	Block 4: Cloning Project	Dec 7	Block 4: Cloning Project
Dec 12	Block 4: Cloning Project	Dec 14	Block 4: Cloning Project Results Presentation
Final Lab Practical Dec 18, 8:00-10:00 & 11:00-1:00

	Activities Skills / Techniques / Concepts	Handout/Readings
BLOCK 1 - BASIC SKILLS
Labs Handounts and links	Links of Intrest : PDB Restriction Enzymes ; NEB Restrction Enzyme Site ;1 KB Ladder Info ; 1KB Lader Instructions Ranin Pipet Specs ; Lab Notebook Grading Rubric ;	Required Book : 1-22; 29-40; 53-59; Labs: Lab 1 Pipetting and Sterile Culture ; Lab 2 Transformation ; Lab 3 Plasmid Purification ; Lab 4 Restriction Digest and Agarose Gel ; Required Links: LabArchives (ELN); Lab Math; Sterile Tech, Design of Bacterial Promoter; addgene plasmid book; Qiagen Miniprep Handbook; SYBR and DNA Ladder; SYBR DNA Gel Stain; Helpful Videos and Links: Pipetting Video;Restriction Digests, Enzymes and Troubleshooting Videos. Transformation Tutorial; Casting an agarose gel; Alkaline Lysis Plasmid Prep ; Qiagen MiniPrep ; The bacterial promotor ;
Lecture Handouts and Assignments	DNA, RNA structure & basics, plasmids; cloning vectors, promoters, operons and genetic gene regulation, restriction enzymes II and IIs, antibiotic mechanisms... Safety Handout; ELN Lab Book ; Dilutions Calculations Practice Set; 8 Steps to Writing an SOP; Effective use of an SOP; USD SOP Template (word file); SOP assignment. Work in your assigned group. Pick an instrument or piece of equipment (email if you aren't sure if your choice is appropriate) google, search, find the instructions and create an SOP based on the links, your course and other background information you find on the web. Use the word form linked here for your assignment. Pipetting and Sterile Culture Basic E. coli Information for Molec Bio; Promoters and Plasmids; Agarose Gels ; Plasmid Purification ; Eton Bioscience - select regular sequencing; USE ONLY SnapGene - for sequence analysis and many other ; pQE60 primer Info ; Sequence (ab1 format - use SnapGene to read) SAM : BLM ; SGAP ; SHKK ; MMB ; FSKCWC ; NGEP ; EAC ; CEJ ; CSJ ; WT ; Here is the pQE60 sequence (word doc) (SnapGene format) and the mature gMDH DNA Sequence ; here is the UniProtKB for P19446 (it includes a precoursor we don't have in this gene)
BLOCK 2 - Bioinformatics, Cloning, Sequencing and PCR
Labs Handounts and links	Links of Interest: See links to the right in this section	Required Book : Chapters 2, 5, 8, and 9 Labs: Lab 1 Bioinformatics and sequence analysis; Lab 2 Field Trip; Lab 3 SNP analysis; GEL Lab 4 qPCR Homework : Cloning Assignment I; Cloning Assignment II; Take Home Exam I ; Exam I answers Required Links:PCR Primer Design I; PCR Primer Design II; PCR for adding restriction sites I; PCR for adding restriction sites II; PCR Primer Design I; PCR Primer Design II; PCR for adding restriction sites I; PCR for adding restriction sites II; Cloning Overview; Advance Site Directed Mutagenesis ; Basic cloning flow : Tips for Cloning ; Cloning into a Plasmid ; To gel or column purify PCR product for cloning ; Helpful Videos: All types of cloning methods ; Golden Gate Cloning ; Primer Design Tutorial I, PCR Design Tutorial II,
Lectures	Cloning Strategies, Basic and Advanced PCR, Sequencing (NextGen sequencing, primer design Deep Seq)
BLOCK 3 - Cloning Stratagies and Project Design
Labs Handounts and links	TBA	Required Book : TBA Labs: TBA Links: Cloning Project Info Coming Soon
Lectures	TBA
BLOCK 4 - Cloning Project
Labs  9-10	Malate Dehydrogenase Cloning/Subcloning Projects - PCDNA Invitrogen Manual - pFLAG tag Manual - HA Map - SYFP2 Turq GFP Tandam Clone paper - mTurq paper - pTaq paper - Novagen pET28 Manual - pET28a map - hCS Info sheet - hCS snapgene map - hMDH1 Info sheet - hMDH1 snapgene map - hMDH2 Info sheet - hMDH2 snapgene map - pQE60 - MGH Seq Info - pGEX4T-1 Snapgene - MGH Snapgene map - - .	Required Book : Open book - use for primary reference Labs: Projects and Milestones ; Project Description ; Initial Visit Planning Sheet ; UPDATED FINAL VERSION OF FINAL REPORT FORMAT/INFORMATION; Links:TBA
Lectures	Cloning, site directed mutagenesis, sequencing, PCR and rtPCR
Presentations and Finals Information
		FINAL EXAM NEW!!!!!!! Molec Tech Exam II 2017.docx