Bio113 Weekly Lab Resources
(100% in person)

A. Malcolm Campbell, Fall 2022 

1 Sep Week 1: getting started, pipets, serial dilutions, GenBio-MAPS if first intro bio course

1) Getting started    

• Lab Manual for Week 1 (read before lab)
• answer questions A - D from the lab manaul

2) Serial Dilutions

• Micropipets explained on paper
• How to use a micropipet (watch before lab; 6:08 video)
• How to use a spectrophotometer (watch before lab; 5:03 video)
• Calculate dilutions (watch before lab; 1:22 video)
• Excel file for calculating series A dilutions
• Series A data (watch before lab; 2:41 video)
• Instructions for generating good Excel graphs (Mac) (Windows)
• Series B & C data (watch before lab; 0:27 video)
• Excel files for series B and series C
• Series D data (watch before lab; 0:25 video)
• Excel file for series D
• Analyze all 4 graphs (watch before lab; 0:24 video)
• Final report template Word file
  

8 Sep Week 2: Boil and Cool oligos (day before lab), Golden Gate Assembly, transformation, plating for version 1

Cloning your experimental promoter (v1)

• Lab Manual Week 2

Build dsDNA from oligos

• Protocol for making dsDNA promoter from 2 oligos
• Mix oligos before boiling and cooling (2:51 video)
• Boiling the oligos (0:35 video)

Golden Gate Assembly (GGA) and Transformation

• GGA protocol after boiling and cooling paired oligos
• Diluting oligos before GGA (3:10 video)
• Start the thermocycler for 20 cycles of GGA (1:19 video)
• Keynote, PPT or PDF presentation on GGA (optimized for Keynote)
• J119137 = pClone Red
• cutting and cloning DNA animation
• easier to read DNA sequences of oligos Excel file (find your 2 oligos)
• oligo full documentation PDF (find your 2 oligos)
• Transforming E. coli (we use strain JM109)
• Transform strain JM109 E. coli cells with GGA (2:10 video)
• Plate transformed strain JM109 E. coli cells onto LB amp agar (100 µg/mL) (2:47 video)
  

15 Sep Week 3: Grow colonies (day before lab), measure phenotype, start genotyping

Pick colonies and grow overnight

• Lab Manual Week 3
• Check colonies after overnight incubation, 37^o C (1:30 video)
• Pick colonies, add to LB amp (100 µg/mL) media (1:32 video)
  

Determine phenotypes and genotypes

• Protocol for reading 96-well plates
• Pipet 200 μL of overnight cultures into 96-well plate (2:13 video)
• protocol for PCR of v1 promoters in J119137
• PCR animation (1:27 video)
• Setup PCR from overnight cultures above (1:33 video)
• Start thermocycler for PCR (1:07 video)

22 Sep Week 4: gel electrophoresis of PCR, prepare for oral presentation

Complete plasmid PCR genotyping

• Lab Manual Week 4
• Protocol for gel electrophoresis
• Load the 2% agarose gel (2:13 video)
• Turn on the current to size-separate DNA (1:03 video)
• Molecular Weight markers

Generate your PPT slides for oral presentations

• Read tips for oral presentations
• Grading rubric for oral presentations

29 Sep Week 5: complete oral presentation

• Lab Manual Week 5
• Read tips for oral presentations
• Grading rubric for oral presentations

6 Oct Week 6: oral presentations and work on V2 promoter designs

Communicating Science to Colleagues

• Lab Manual Week 6
• Give oral presentation and feedback for other 3 groups
• iBOP Bingo for oral presentations
• Skim the one paper describing your promoter
• entP (blue)
• pbp1b (green)
• glp (red)
• hutP (yellow)
• oligator web tool for making dsDNA used for GGA (your instructor can help you with this web tool)
• Research Proposal Template (will be graded)

13 Oct Week 7: Golden Gate Assembly, transformation, plating (v2)

Cloning your experimental promoter (v2)

• Lab Manual Week 7

Build dsDNA from oligos (v2)

• Protocol for making dsDNA promoter from 2 oligos
• Making dsDNA from two oligos (1:19 video recap)

Golden Gate Assembly (GGA) and Transformation (v2)

• GGA protocol after boiling and cooling paired oligos
• Starting GGA (1:43 video recap)
• J119137 = pClone Red
• cutting and cloning DNA animation (refresher)
• v2 oligos Excel format (find your 2 oligos)
• v2 oligos, full documentation PDF (find your 2 oligos)
• Transforming E. coli (we use strain JM109)
• Transform and plate JM109 E. coli cells with GGA (1:30 video recap)

20 Oct Week 8: Pick colonies, grow in LB amp  (v2), miniprep plasmid DNA

Pick colonies v2 + colony X1 from v1 promoter

• Lab Manual Week 8
• Pick colonies, start overnight cultures (1:02 video recap)

Determine phenotypes and genotypes

• Protocol for reading 96-well plates 
• Pipet 200 μL of overnight cultures into 96-well plate (2:13 video)
• Protocol for mini-prepping pladmid DNA
• miniprep setup (1:17 video)

27 Oct Week 9: sequence plasmid DNA

• Lab Manual Week 9
• DNA dideoxy sequencing method (5 min video)
• Protocol for sequencing DNA
• Protocol for nanodropping DNA to quantify concentration
• Dilution calculation
• Excel template for sequencing reactions
• mix plasmid DNA with sequencing primer (2:34 video)

Send off DNA to be sequenced

3 Nov Week 10: analyze DNA sequences

compare expected sequence to sequencing results

• Lab Manual Week 10
• verbal instructions for DNA analysis (0:51 video)
• Clutal omega sequence alignment site
• Clustal omega submission template
• download morning sequencing data
• download afternoon sequencing data

10 Nov Week 11: Final Lab Report due next week

Prepare your final lab report

• Lab Manual Week 11
• Modify this lab report template

17 Nov Week 12: Finish lab report

Submit Final Lab Report

• email your final lab report to Dr. C. He will reply to confirm submission.
• Final lab report template (Word file)

8 Dec Week 13: graduate fellowships, course evaluations, GenBio-MAPS if last college bio course

Last lab meeting

• Watch slide show about fellowships that pay you to travel
• Final course evaluation
• GenBio-MAPS if this is your last intro bio course at Davidson
  

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