Week 1 (0427-0429)

1. Prepare competent BL21 cells with calcium chloride
2. Make LB plates
3. Transformation of GFP
4. Pick colony

Detail

1. Make LB broth (400 ml) and LB agar (400 ml)
2. GFP and RFP transformation practices



3. Cell preparation (BL21) competent cell using CaCl2
4. Make LB agar plate
5. Transformation of GFP
6. Pick colony (concentration: 1000, 500, 100)
7. Incubate at PuiChing: cover the tube with the cap at second stop (fully covered, tight), use parafilm to pack the tubes. Make sure that all tubes are vertical.

Week 2 (0504-0507)

1. Check GFP and OD for growth curve
2. Pick colony of GFP

Week 3 (0514-0521)

1. Check hydroponic pH and conductivity
2. Check OD and GFP for growth curve
3. Renew nutrient solution of hydroponic system

Detail

Week 4 (0523-0528)

1. Check hydroponic pH and conductivity
2. make plates
3. Transformation of GFP
4. pick colony of GFP
5. glycerol stock of GFP

Week 5 (0530-0601)

1. Check hydroponic pH and conductivity
2. Make LB agar and broth, check ABS and FL for growth curve

Detail

Week 6 (0609-0610)

1. Transformation of pH shooting (twice)
2. Make LB agar
3. Spread BL21 plates

Detail

Week 7 (0617)

1. Transformation of glsA, pH shooting and sf-gfp
   result: only glsA has colonies

Detail

Week 8 (0803-0805)

1. Primer design for PYL8 and T2R4
2. Make LB agar, broth, and plates
3. Spread BL21 cells
4. Pick three colonies, and prepare competent cells with calcium chloride

Detail

Week 9 (0808-0812)

1. Transformation of glsA, K, pH shooting and control
2. result: only glsA and control had colonies, so we will midi K and pH shooting
3. Pick colony and culture glsA and control
4. Make pH 4, 5, 6, 7 LB broth for functional test
5. 1st time glsA and control functional test
6. Transformation of K and pH shooting with DH5a cells
7. Pick colony and culture
8. Primer design for glsA and ldhA

Detail

Week 10 (0815-0819)

1. 2nd time glsA and control functional test: measure pH and OD
2. result: likely contaminated so we do control again
3. Midi prep of K and pH shooting
4. Transformation of glsA, K, pH shooting, and control
5. Pick colony and culture control
6. Control functional test

Detail

Week 11 (0822-0826)

1. Make pH 8, 9, 9.5 LB broth, autoclave for functional tests
2. culture control, pH shooting, spread glsA and K plate
3. functional test of pH shooting and control: measure pH, OD and FL
4. Pick colony and culture glsA and K
5. functional test of K and glsA

Detail

Week 12 (0829-0901)

1. Make pH 5, 6, 7 LB broth, autoclave for functional tests
2. functional test of pH shooting and control: measure pH, OD and FL
3. spread glycerol stock cells (pH shooting and control)
4. Pick colony and culture

Week 13 (0905-0908)

1. Western Blot of pHS and glsA
2. results: no visible bands, too little sample
3. pick colony
4. mini prep PYL8 and T2R4
5. measure growth of soybean and water spinach in different pH nutrient solution

Detail

Week 14 (0914-0918)

1. Design and start functional tests for hormone system
2. measure growth of soybean and water spinach in different pH nutrient solution
3. Redo pH system western blot with 20 microliter samples
4. result: success
5. Prepare cDNA for RT-qPCR for PYL8 and T2R4 genes

Detail

Week 15 (0920-0924)

1. RT-qPCR glsA and ldhA
2. Culture and extract protein for Western blot of PYL8 and NSP4-T2R4
3. Transformation of new pH system vectors using BL21 and DH5a cells (low efficiency as it is not IPTG induced)
4. Pick colony (used wrong antibiotics)

Detail

Week 16 (0926-0928)

1. RT-qPCR for PYL8 and T2R4
2. results:
1. PYL8: 1.33 fold change against BL21
2. T2R4: 1.60 fold change against BL21
3. Transformation of new pSB1C3 vectors
4. Western blot for PYL8 and NSP4-T2R4
5. result: Obvious band for PYL8, no band for NSP4-T2R4
6. Start functional tests of new pSB1C3 vectors using raspberry pi and cloud system

Detail