Bio113 Weekly Lab Resources
(100% in person)

2) Serial Dilutions

• Micropipets explained on paper
• How to use a micropipet (watch before lab; 6:08 video)
• Calculate dilutions (watch before lab; 1:22 video)
• Excel file for calculating serieal dilutions
• Series A data (watch before lab; 2:41 video)

Jan 23-24 Week 2: serial dilutions, plate reader, graphing data

1) Before Lab

• Lab Manual for Week 2 (read before lab)
• answer questions A - D from the lab manaul

2) Evaluating Serial Dilutions

• Generate your own data for 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
  

Jan 30-31, Week 3: Boil and Cool oligos (Monday 4:30), Golden Gate Assembly, transformation, plating for version 1 (v1)

1) Cloning your experimental promoter (v1)

• Lab Manual Week 3

2) Build dsDNA from oligos

• Protocol for making dsDNA promoter from 2 oligos (start with step 3#)
• Mix oligos before boiling and cooling (2:51 video)
• Boiling the oligos (0:35 video)

3) 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)
• PPT or PDF presentation on GGA (optimized for Keynote)
• J119137 = pClone Red
• cutting and cloning DNA animation (1:12 video)
• 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)
  

Feb 6-7, Week 4: Grow colonies (Monday 4:30), measure phenotype, start genotyping

1) Pick colonies and grow overnight

• Lab Manual Week 4
• Check colonies after overnight incubation, 37^o C (1:30 video)
• Pick colonies, add to LB amp (100 µg/mL) media (1:32 video)
  

2) Determine phenotypes and genotypes

• 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)
• Protocol for making glycerol stocks
• Protocol for reading 96-well plates
• Pipet 200 μL of overnight cultures into 96-well plate (2:13 video)

Feb 13-14, Week 5: gel electrophoresis of PCR, prepare for oral presentation

1) Complete plasmid PCR genotyping

• Lab Manual Week 5
• 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

2) Generate your PPT slides (not Google) for oral presentations

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

• Lab Manual Week 6
• Oligator: generate top and bottom strands for GGA cloning
• Read tips for oral presentations
• Grading rubric for oral presentations

Feb 27-28, Week 7: oral presentations and work on V2 promoter designs

Communicating Science to Colleagues

• Lab Manual Week 7
• Give oral presentation and feedback for other 3 groups
• iBOP Bingo for oral presentations

Prepare Research Proposal for v2 Promoter

• entB (blue)
• pbp1b (green)
• glp (red)
• hutP (yellow)

Mar 12-13 Week 8: make dsDNA (Monday 4:30), GGA, transformation, plating (v2) 

Cloning your experimental promoter (v2)

• Lab Manual Week 8

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 (1:12 video recap) (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)

Mar 19-20 Week 9: Pick colonies (Monday 4:30), grow in LB amp  (v2), miniprep plasmid DNA

Pick colonies v2 + colony X1 from v1 promoter

• Lab Manual Week 9
• 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 (1:17 recap video)
• Protocol for mini-prepping pladmid DNA
• miniprep setup (1:17 video)

Mar 26-27 Week 10: sequence plasmid DNA

• Lab Manual Week 10
• 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

Apr 2-3 Week 11: analyze DNA sequences

compare expected sequence to sequencing results

• Lab Manual Week 11
• verbal instructions for DNA analysis (0:51 video)
• Kalign sequence alignment site
• Clustal omega submission template
• download Tuesday sequencing data
• download Wednesday sequencing data

Apr 9-10 Week 12: Final Lab Report due next week

Prepare your final lab report

• Lab Manual Week 12
• Modify this lab report template

Apr 16-17 Week 13: Finish lab report

Submit Final Lab Report

• Lab Manual Week 13
• email your final lab report to Dr. C. He will reply to confirm submission.
• Final lab report template (Word file)

Apr 23-24 Week 14: graduate fellowships, course evaluations

Last lab meeting

• Watch slide show about fellowships that pay you to travel
• Final course evaluation

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