Notebook

February

    Molecular biology class
    E. coli genomic DNA preparation
    E. coli transformation

March

    Instrument operation

April

    Project brainstorming: Product Positioning, Monolignol proportion

May

    Culture selection: compare Yeast, E. coli, Acetobacter aceti
    Gene design

June

    Interlab experiment: Calibration 1, 2, 3

July

  7/3
    Start Interlab experiment: cell measurement
  7/5
    Yeast Extract–Peptone–Dextrose (YPD) formulation
  7/9
    Yeast (X33) culture
  7/13
    Fundraising briefing session
  7/16
    Communicate with NCKU (Interlab & project)
  7/18
    Communicate with BIT (project)

August

8/5~8/12

    Cell growth, sampling, and assay
    Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI
    Transformation pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18 (15µl) into 20µl ECOS™
    101 Competent Cells [DH5a]
    Ligation pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18
    Digest pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI

8/13~8/19

    Miniprep pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18
    Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI
    Digest pGAPZ A with SalI
    Incubate with DNA Polymerase I, Large (Klenow) Fragment remove SalI restriction site
    Ligation pGAPZ A-SalI(pGAPZ A-1)
    Transformation of pGAPZ A-SalI(pGAPZ A-1)
    Minipreparation of pGAPZ A-SalI(pGAPZ A-1)

8/20~8/26

  Gel extract
    Digestion – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)
    Ligation - pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18

8/27~9/2

    Digestion – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)
    Ligation – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)

    Transform
      1. pGAPZ A-1 + HIS4 + Px16
      2. pGAPZ A-1 + HIS4 + Px18
      3. pGAPZ A-1 + HIS4 + Lac1
    Digest pGAPZ A HIS4(X2) with AgeI, EcoRI
    pGAPZ A_Px16, pGAPZ A_Px18 , pGAPZ A_Lac1 Traditional plasmid DNA extraction
    Yeast SMD1168 Transformation(pGAPZ A_Px16, pGAPZ A_Px18 , pGAPZ A_Lac1)

September

9/3~9/9

    PCR—pGAPZ A-1 + HIS4 + Px16, Px18, Lac1
    miniprep – pGAPZ A-1 + HIS4 + Px16, Px18
    Transform – pGAPZ A-1 + HIS4 + Lac1
    Digestion – pGAPZ A-1 + HIS4+lac1

9/12~9/14

    Colony PCR gel electrophoresis check - HIS4+Px16, HIS4+Lac1
    Culture HIS4+Px16, HIS4+Lac1
    Colony PCR gel electrophoresis check - HIS4+Px16, HIS4+Lac1
    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 colony PCR
    Plasmid miniprep - HIS4+Lac1, HIS4+Px16

9/24~9/27

    Culture HIS4+Lac1 x3
    Prepare yeast competent cells (SMD1168)
    Traditional plasmid DNA extraction Day1
    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 colony PCR
    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 Western blot
    Prepare YPD plates
    Prepare the amino acids of SD plates
    Traditional plasmid DNA extraction Day2
    Plasmids contain HIS4 cut with SalI
    Yeast transformation - Px16, Px18, Lac1 (plasmids contain HIS4)

October

10/8~10/11

    Prepare yeast competent cells (SMD1168)
    Colony PCR (HIS4+Px16)
    Safety (30℃, 50℃, 70℃)
    Yeast transformation - Px16, Lac1 (plasmids contain HIS4)

    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 Western blot