Kelsey Lawton Environmental Genomics, Spring 2021 1-30-21 Obtained soil samples from Northern NY with Coordinates 44.064009, -75.481864 2-01-21 Placed 1g of soil into 5ml of TB and incubated at 33C in a shaker for about 1 week 2-06-21 Pipetted 100ul of the LB with soil into a tube of fresh LB and incubated at 35C for 1 week 2-19-21 Pipetted 100ul of the previous culture into 4 different tubes and incubated at 30C and 200 rpm : 1. 5ml CSM + 5ul PX 2. 5ml SSM + 5ul PX 3. 5ml CSM + 5ul MP 4. 5ml CSM + 5ul MP 03-03-21 Realized that we used the wrong tube of Pesticides (too high of a conc.)so these will need to be redone. 03-05-21 Pipetted 100ul of the previous culture (in LB) into 2 different tubes and incubated at 30C and 200rpm: 1. 5ml CSM + 5ul PX 2. 5ml SSM + 5ul PX These were done separately because we were out of MP. 03-09-21 Pipetted 100ul of the previous culture (in LB) into 2 different tubes and incubated at 30 degrees and 200rpm: 1. 5ml CSM + 5ul MP 2. 5ml SSM + 5ul MP 03-15-21 Pipetted 200ul of the previous culture (week 1) into new tubes with minimal media and pesticides as follows: 1. 5ml CSM + 5ul PX 2. 5ml SSM + 5ul PX 3. 5ml CSM + 5ul MP 4. 5ml CSM + 5ul MP These were incubated at 30C and 200rpm 03-22-21 spread 100ul of PX on two plates and 100ul of MP on two plates and let dry overnight in the fridge. Added 1 ml of week 2 samples in minimal media and pesticides into 4ml of LB to grow overnight. 03-23-21 streaked the samples in LB onto the corresponding plates and incubated at 37C Pipetted 200ul of the previous culture (week 2) into new tubes with minimal media and pesticides as follows: 1. 5ml CSM + 5ul PX 2. 5ml SSM + 5ul PX 3. 5ml CSM + 5ul MP 4. 5ml CSM + 5ul MP 04-01-21 spread 100ul of PX on two plates and 100ul of MP on two plates and let dry overnight in the fridge to make grid plates. 04-02-21Drew grids on the 4 plates that dried overnight, and numbered the boxes. Picked colonies from each plate from 03-23-21 and dotted them into each box on the corresponding grid plate and incubated them at 37C. The non grid plates were parafilmed and stored in the fridge. Pipetted 200ul of week 3 into new tubes with minimal media and pesticides as follows: 1. 5ml CSM + 5ul PX 2. 5ml SSM + 5ul PX 3. 5ml CSM + 5ul MP 4. 5ml CSM + 5ul MP 04-06-21 took grid plates from 04-02-21 out of the incubator, parafilmed them and stored them in the fridge. 04-12-21Took week 4 tubes out of the incubator and stored it in the fridge. 04-16-21 Pipetted 200ul of week 4 into new tubes with minimal media and pesticides as follows: 1. 5ml CSM + 5ul PX 2. 5ml SSM + 5ul PX 3. 5ml CSM + 5ul MP 4. 5ml CSM + 5ul MP 04-21-21Placed 1ml of the week 5 samples into 4ml of LB and let them grow for 6hrs. Spread 100ul of PX on two plates and 100ul of MP on two plates and let them dry for 6hrs. Streaked the samples in the LB onto the corresponding plate and placed in an incubator at 37C 04-22-21 Spread 100ul of PX on two plates and 100ul of MP on two plates and let dry overnight. Drew grids on the 4 plates that dried overnight, and numbered the boxes. Picked colonies from each corresponding plate from 04-22-21and dotted them into each box on the corresponding grid plate and incubated them at 37C. 04-23-21 lysed the colonies to prep them for PCR by placing a small amount of each colonies using a toothpick in 100ul of sterile water and heating them to 95C for 5 minutes. There were 30 samples total. Below is the code for the labels. Week 2 Samples A- CSM + MP B – CSM + PX C – SSM + MP D – SSM + PX Week 5 Samples E – CSM + MP F – CSM + PX G – SSM + MP H – SSM + PX The number on the tube corresponds to the number on a single grid plate Samples that were Sequenced KL A1 KL A2 KL A3 KL A4 KL A5 KL A6 KL A7 KL A8 KL A9 KL B1 KL B2 KL B3 KL B4 KL B5 KL B6 KL C1 KL C2 KL C3 KL D4 KL D5 KL E1 KL F1 KL F3 KL G1 KL G2 KL G3 KL H1 KL H2 KL H3 KL H4
