Team:Munich/Notebook

Notebook

Assay-Team

April 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
1	2	3	4	5	6	7
8	9	10	11	12	13	14
15	16	17	18	19	20	21
22	23	24	25	26	27	28
29 seeding	30 test PEI conditions

May 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
		1 medium exchange	2 exosome precipitation	3 exosome precipitation	4	5
6	7	8 SDS-PAGE (exosomes)	9 silver staining	10	11	12
13	14	15 DLS	16 exosome precipitation, SDS-PAGE (exosomes)	17 exosome precipitation, SDS-PAGE (exosomes)	18	19
20 DLS	21	22	23	24 RNA isolation exosomes (cells)	25	26
27 seeding	28 RNA isolation	29 exosome precipitation	30 exosome precipitation	31 SDS-PAGE

June 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
					1	2
3 seeding	4 transfection	5 harvest	6 exosome precipitation	7 exosome precipitation, SDS-PAGE	8	9
10	11 DLS	12 in vitro transcription of fluc-MS2	13 His-purification Exosomes	14 SDS-PAGE	15	16
17 in vitro transcription of fluc-MS2	18 in vitro transcription of fluc-MS2	19 in vitro transcription	20	21 RNA isolation, cDNA synthesis, qPCR	22	23
24	25 RNA isolation (RNeasy Mini kit vs. Trizol)	26 qPCR, Western Blot	27	28	29	30

July 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
1 in vitro transcription of fluc-MS2	2 PCR, agarose gel	3	4 His-Purification Exosomes & VLP, Lysis VLP & Exosomes, RNA isolation, Phenol-/Chloroform extraction	5	6	7
8 DNA digestion	9 reverse transcription	10	11 agarose gel	12	13	14
15	16 Western Blot; DNAse treatment VLPs	17 SDS-PAGE; Western Blot	18 Western Blot	19 non-reducing and native PAGE	20	21 SDS-PAGE
22 reverse transcription	23 qPCR	24 agarose gel	25 reverse transcription	26 Western Blot	27	28
29 Western Blot native; Exosome preciptiation	30 Exosome precipitation, RNA isolation, Western Blot	31 Western Blot

August 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
			1 Western Blot	2 Western Blot; RNA isolation VLPs	3	4
5 non-reducing and reducing PAGE	6 Lysis of VLP samples, DNAse treatment, RNA isolation	7 reverse transcription, agarose gel	8 Exosome preciptiation, RNA extraction, qPCR Exosomes and VLPs	9 Exosome precipitation, RNA extraction, agarose gel	10	11
12 Western Blot	13 Western Blot	14 VLPs RNA isolation (Trizol)	15 Exosomes RNA isolation (Trizol)	16 SDS-PAGE; Western Blot; VLPs Trizol	17	18
19 SDS-PAGE; Western Blot; reverse transcription of Exosomes	20 PCR; Western Blot; reverse transcription of Exosomes	21 Western Blot; reverse transcription of Exosomes; Dpnr1; qPCR	22	23 qPCR VLPs and Exosomes; Western Blot	24	25
26 RNA isolation	27 Western Blot; reverse transcription of VLPs; VLPs RNA isolation	28 reverse transcription Exosomes and VLPs; Western Blot; RNA isolation Exosomes; qPCR	29 Western Blot	30 qPCR Exosomes and VLPs	31

September 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
						1
2 rRNA isolation	3 DNase treatment; RNA isolation Trizol; reverse transcription	4 RT	5 SDS-PAGE; Western Blot	6 Western Blot; RNA extraction	7	8
9 Rt, SDS-PAGE, Western Blot	10 Western Blot	11 RIPA buffer, Western Blot	12 RNA extraction, Western Blot	13 RNA extraction, SDS-PAGE, Western Blot	14 RNA extraction	15 RNA extraction
16 RNA extraction; qPCR	17 RT, qPCR	18 qPCR	19 Agarose gel; RNA extraction; RT	20 RNA extraction; RT	21 RNA extraction; RT	22
23 RT, qPCR, Western Blot	24 Western Blot	25 RNA extraction, RT, Western Blot	26 Western Blot	27 PCR, RNA extraction, RT	28 RT, RNA extraction	29 RNA extraction, RT
30 RNA extraction, RT, Western Blot

October 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
	1 RT, Western Blot	2 qPCR	3 Western Blot	4 qPCR, DLS, Western Blot	5	6
7 Western Blot	8 Western Blot	9 RNA extraction	10 RNA extraction; RT	11 RNA extraction	12 RNA extraction	13 RNA extraction
14 qPCR	15 qPCR	16	17 qPCR	18 Western Blot	19 Western Blot	20
21	22	23	24	25	26	27
28	29	30	31

Exosome-Team

April 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
1	2	3	4	5	6	7
8	9	10	11	12	13	14
15	16	17	18	19	20	21
22	23	24	25 Fussenegger plasmids	26 Fussenegger plasmids	27	28
29 seeding	30 transfection

May 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
		1 medium exchange	2 exosome precipitation	3 seeding	4	5
6 seeding	7 transfection booster blasmid	8 harvesting	9	10	11	12
13 seeding	14 transfection booster plasmid	15 medium exchange	16 harvesting	17 splitting	18	19
20 splitting	21	22 cloning	23 cloning	24 cloning	25	26
27 cloning	28 cloning	29 cloning	30	31 splitting, cloning

June 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
					1	2
3 cloning E10, E11; seeding	4 cloning E10, E11; transfectin E6 + E12	5 cloning E13	6 cloning E13	7 cloning E13	8	9
10 seeding	11 transfection	12 cloning E10, E11	13 cloning E10, E11	14 cloning E10, E11	15	16
17	18 seeding	19 seeding, transfection	20 transfection for HiBiT + qPCR	21 HiBiT	22 splitting	23
24 splitting	25 cloning E9/V15	26	27 splitting	28	29	30

July 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
1	2 transfection, seeding	3 transfection	4 lysis screening	5 HiBiT	6	7
8 seeding	9 transfection (reuptake assay)	10 reuptake assay (medium transfer)	11 reuptake assay, HiBiT, His-Tag purification	12 seeding, reuptake assay	13 transfection (His-purification and reuptake assay)	14
15 seeding and splitting	16 transfection	17	18 reuptake assay, His-Tag purification	19 splitting, reuptake assay	20	21
22 splitting	23	24 cloning	25 seeding and splitting, cloning	26 seeding, reuptake assay	27	28
29 splitting and seeding	30 reuptake assay	31 reuptake assay

August 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
			1	2 splitting	3	4
5 seeding	6 cloning E16, transfection (viability assay, qPCR)	7 cloning E16	8 cloning E16; viability assay (trypan blue)	9	10	11
12 seeding	13 cloning: E17; transfection (qPCR, purification)	14	15 reuptake assay, cloning, seeding, HiBiT	16 transfection, seeding, reuptake assay	17 seeding, reuptake assay	18 transfection, seeding, His-Tag Purification
19 reuptake assay (medium transfer); seeding; cloning	20 His-Tag purification, seeding, transfection, cloning	21 reuptake assay, LDH assay	22 His-Tag purification	23	24 transfection	25
26 seeding	27 transfection	28 cytotoxicity LDH assay	29 cytotoxicity LDH assay (medium transfer), His-Tag Purification	30 cytotoxicity LDH assay, His-Tag purification	31

September 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
						1
2 seeding	3 transfection	4 seeding recipient cells (cytotoxicity LDH assay)	5 cytotoxicity LDH assay	6 His-Tag purification, LDH assay	7	8
9 cloning	10 transfection	11 cytotoxicity LDH assay	12 EVOS, cloning	13 EVOS, cloning, transfection, splitting	14 EVOS	15
16 seeding, HiBiT	17 transfection (cytotoxicity LDH assay)	18 LDH assay	19 LDH assay, cloning, splitting	20 LDH assay, HiBiT	21	22
23 splitting, cloning	24 splitting and seeding	25 transfection	26 splitting and seeding	27 transfection	28 splitting and seeding	29 reuptake assay
30 reuptake assay, splitting and seeding

October 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
	1 transfection (HiBit 96-well plate, purification)	2 cloning	3 HiBit	4 His-Tag purification, cloning, reuptake assay	5	6 seeding
7 transfection	8 cloning, seeding	9 His-Tag purification, transfection, HiBiT	10 LDH assay, cloning, HiBiT	11 LDH assay, HiBiT, splitting and seeding	12 LDH assay	13
14	15	16 His-Tag purification	17	18	19	20 HiBiT
21	22	23	24	25	26	27
28	29	30	31

VLP-Team

April 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
1	2	3	4	5	6	7
8	9	10	11	12	13	14
15	16	17	18	19	20	21
22	23	24	25	26	27	28
29	30

May 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
		1	2	3	4	5
6	7	8	9	10	11	12
13	14	15	16	17	18	19
20	21	22	23	24	25	26
27	28	29	30	31

June 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
					1	2 splitting
3 splitting and seeding, plasmids from Gruber	4 transfection	5 cloning V1 + V4 + V5 + V8	6 HiBiT, cloning V1 + V4 + V5 + V8	7 splitting, cloning V1 + V4 + V5	8	9
10 cloning V1 - V8	11 cloning V1 - V8	12 cloning V8	13 seeding	14 transfection	15	16
17 cloning	18 cloning V8 - V10	19 cloning V9 + V10	20 splitting and seeding, cloning	21 cloning V12 + V13, transfection	22 cloning	23 cloning V10 + V14 + V16 + V9
24 cloning V10 + V14 + V16, splitting	25 harvesting	26 cloning V10 + V14 + V16	27 HiBiT, splitting	28 cloning V11, splitting	29 cloning V11	30 cloning V10 + V11 + V14, transfection

July 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
1 cloning V15, V25-V28, seeding and splitting	2 cloning V15, V17, V25-V30, transfection	3 harvesting, cloning V28, V29, V15, V17	4 medium exchange, harvesting, cloning V15 + V30	5 cloning V25 - V28, HiBiT	6 HiBiT	7
8 splitting	9 cloning V15	10 cloning V15	11 HiBiT assay (triton test), splitting	12	13	14 splitting and seeding
15 transfection (HiBiT, exosomes for lysis test)	16	17 medium exchange, HiBiT calibration curve	18 HiBiT, splitting, cloning	19 splitting	20	21 cloning
22 seeding (bioinfo), cloning V31 + V32	23 seeding, cloning V31 + V32	24 transfection, cloning V31 + V32	25 medium exchange	26 cloning V31 + V32, splitting, fluorescence microscopy	27 HiBiT	28
29 seeding (qPCR, HiBiT, purification)	30 transfection (purification, HiBiT, qPCR)	31

August 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
			1 medium exchange	2 HiBiT 96-well plate, purification (biotin)	3	4
5 seeding (purification, hArc)	6 transfection (purification, hArc)	7	8 medium exchange	9 HiBiT hArc, purification (His, biotin)	10	11
12 seeding (qPCR, purification, HiBiT)	13 transfection (qPCR, purification, HiBiT)	14	15 medium exchange	16 HiBiT (96-well plate), purification (His, biotin)	17	18
19 seeding (bioinfo, reuptake assay, qPCR, purification)	20 transfection (reuptake assay, qPCR, purification)	21 medium exchange	22 reuptake assay	23 biotin purification, reuptake assay	24 reuptake assay	25
26 medium exchange T75, cloning V33, biotin purificatoin	27 cloning V33, bioitn purification	28	29 cloning V33	30	31

September 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
						1
2 seeding for purification and qPCR	3 transfection	4 medium exchange (qPCR)	5 medium exchange (purification)	6 VLP purification (His, Biotin) (1 h incubation)	7 purification (His, biotin) (o/n incubation)	8
9 seeding for purification and qPCR	10 transfection, cloning V34	11 medium exchange (qPCR), cloning V35-V41, E19	12 seeding, cloning V35-V41, E19	13 cloning V35-V41; E19	14	15 cloning V35-V41; E19
16 seeding for purification, qPCR, biotin, cloning V35-V41; E19	17 transfection	18 medium exchange	19 medium exchange (purification)	20 HiBiT, biotin purification, heparin purification	21 SDS-PAGE	22
23 seeding for new constructs (new coiled-coil, fusionconstruct)	24 transfection	25 medium exchange	26 seeding for reuptake assay	27 TEM, HiBiT, heparin purification	28	29 splitting and seeding
30

October 2019

Monday	Tuesday	Wednesday	Thursday	Friday	Saturday	Sunday
	1	2	3	4 seeding	5 transfectin (qPCR)	6 splitting and seeding (purification, cell dilution series)
7 transfectin (purification, cell dilution series)	8 cloning V44 + V45	9 medium exchange, transfectin (RNA secretion over time)	10 HiBiT; heparin purification	11 seeding, DLS	12 transfection (L7Ae specificity check)	13 seeding
14 transfection (purification)	15	16	17 heparin purification	18	19	20
21	22	23	24	25	26	27
28	29	30	31

Characterization

September & October 2019

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