IGEM team meeting notes 5/06/14

Various intros by lots of people present. Unfortunately no notes for introductions. 22 people present. - Registrations on http://www.meetup.com/BioCurious/events/180642612/ and http://www.meetup.com/Counter-Culture-Labs/events/174105062/ - ccl-igem@googlegroups.com - wiki: https://www.counterculturelabs.org/index.php/Vegan_cheese - sci-hub.org

• Patrik summarizes the vegan cheese project and iGEM for new people.

Agenda

Intro

Patrik

What is Casomorphin? Casomorphins are peptides, i.e., protein fragments, derived from the digestion of milk protein casein. (See Beta-casein.)

What are their effects?

Do they need to be considered in the formulation of cheese?

How can people sign up for the iGEM list/stay in touch with the team?

Google Groups Mailing List: ccl-igem@googlegroups.com

Sign up at https://groups.google.com/forum/#!forum/ccl-igem

What is the work that needs to be done by this team?

• Fundraising
  • Sub-team to take the lead on this? Crowdfunding? Who are we reaching out to?
  • Support from corporations? (Mallory can reach out to - All tech in Ky bio ag company, New Harvest, Maria can contact Barracuda Networks) EPA?
  • Fundraising goals $3500 fee for registration, materials $5000 Goal low $15k, medium $25k, high $50k
  • Ask what crowdfunding we are/will do, crowdtilt has been done, going to restart
  • Treasuer to keep track of funds, expenses, and goals with an estimated budget
  • Are donation tax deductible? who's 501c3 would do this? 501c3 applied for though CCL
  • Teach a class related to the project, all funds going to the project
  • Speaking at events i.e. First Fridays
  • go out to speaking events like first Oakland and talk about project
  • Scrappy vegan biohaker mascot - farmer milking a yeast cell with udders for marketing
  • organize a GMO conference!

• Molecular Biology
  • synthetic biology services: IDT http://eu.idtdna.com/pages/landing/igem
    • 25% off an order, up to $800
    • 40% off gBlocks® Gene Fragments, MiniGenes, or Genes
  • What constructs do we want to make?
    • triple-gene, inducible, construct with promoter-regulated transcription of cheese components.
    • single constructs that serve as controls +/- expression tags for purification.
    • kinases needed for the polymerization of Caseino-peptides.
  • Kinases and glycosylation - Do we want to co-express anything?
  • can we create a construct/work around yes/no flow chart. IE: if construct is created and secretes, then...

• Cheese Making
  • what is the process? Is is the same for synthetic cheese when compared to dairy-derived cheese?
  • What are our equipment needs?
  • Experiments?
    • determining the source of fat

Experiments:

• Cheese Track
  • Make powdered casein and make cheese from it
  • powdered casein with Ca2+ and PO4 to make cheese
  • Synthetic Casein + CA2+ to cheese
  • synthetic milk to casein

• Protein Track
  • Express various caseings
  • evaluate mixtures of synthetic and natural caseins
  • optimize casein phophates
  • results in syntehtic milk -- make cheese from syntehtic milk

• Recipe Track
  • Evaluate pH History of Water/Pwder Mixture
  • Develop recipe for reconstituted milk
  • Develop Recipe for Micelle formation
  • Develop recipe for cleaving enzyme
  • artificial micelles from fat emulsifications.

• Background research
  • Cost of productions
  • Are human casains appropriate for adult humans? research about this

$83 in donations tonight (Patrik) - where do those funds go? Who holds them?

Breakout groups

Fundraising

• Maria
• Mallory
• Patrick

Molecular biology

• Brian
• Dimitri
• Teddy
• Johan
• Marc
• Craig R.
• Nikola
• Colin
• Advait

Cheese Making

• Teddy
• Maria
• Brian
• Della

Background

• Dimitri
• Mallory

Molecular Biology

Genes

• Full-length bovine κ Casein
  • B genetic variant AAQ87923.1

• Full-length bovine αs1 Casein
• Full-length bovine αs2 Casein
• Full-length bovine β Casein
  • A2 genetic variant for health benefit
  • B genetic variant for better coagulation

• Full-length human κ Casein P07498.3
• Full-length human αs1 Casein
• Full-length human αs2a Casein CSN1S2A
• Full-length human αs2b Casein CSN1S2B
• Full-length human β Casein
• human caseinomacropeptide
• FAM20C Kinase

Promoters

3 promoters (low, mid, high)

Selection Markers

http://protocols.mmml.nl/protocols/yeast/yeast_selectable_markers.php

OriC

Purification Tags

Constructs

• Full length single peptides
• do promoters as separate biobricks (if needed)
• have other iGEM projects done protein secretions

HIGH Copy Vector

    Low Promoter

    SS+

    Mid Promoter

    SS+

    TAG

    High Promoter

    SS+

MID Copy Vector

    Low Promoter

    SS+

    Mid Promoter

    SS+

    TAG

    High Promoter

    SS+

LOW Copy Vector

    Low Promoter

    SS+

    Mid Promoter

    SS+

    TAG

    High Promoter

    SS+

Database of constructs, sequences and restricitons

HiFi Polymerase - PfuUltra