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miRNA Detection

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Experiment #1

Attempt #1

GOAL: Test the strength of L7ae

3

complete

7/14/14: Seed cells

22

complete

7/15/14: Transfect MAV1212-hEF1a-eBFP2 (1084.0 ng/uL), Mutant hEF1a-2xKturn-eGFP (551.4 ng/uL) and Wild Type hEF1a-2xKturn-eGFP (287.0 ng/uL) into HEK cells

0

incomplete

7/17/14 (2 pm): Flow Cytometer

DNA Dilution: 5 uL MAV1212-hEF1a-eBFP2, 10 uL Mutant Hef1a-2xKturn-eGFP, 20 uL Wild Type Hef1a-2xKturn-eGFP

24-WELL PLATE TRANSFECTION MAP:

(B)1

MAV1212-hEF1a-eBFP2

(B)₂

MAV1212-hEF1a-eBFP2

//////

(G_M)1

Mutant Hef1a-2xKturn-eGFP

(G_M)₂

Mutant Hef1a-2xKturn-eGFP

(G_M+ B)₁

Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_M+ B)₂

Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_M+ B + L)₁

Mutant Hef1a-2xKturn-eGFP
Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_M+ B + L)₂

Mutant Hef1a-2xKturn-eGFP
Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_WT)1

Wild Type Hef1a-2xKturn-eGFP

(G_WT)₂

Wild Type Hef1a-2xKturn-eGFP

(G_WT+ B)₁

Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_WT+ B)₂

Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_WT+ B + L)₁

Wild Type Hef1a-2xKturn-eGFP
Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_WT+ B + L)₂

Wild Type Hef1a-2xKturn-eGFP
Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

Neg Ctrl

Dummy DNA
With Lipofectamine

Neg Ctrl

Dummy DNA
With Lipofectamine

Neg/Neg Ctrl

No DNA
No Lipofectamin

Neg/Neg Ctrl

No DNA
No Lipofectamine

//////

RESULT: Brief description of experimental outcome (mean transfection efficiency and implications). Link to results page with data analysis.

Attempt #2

2

complete

7/16/14: Seed cells

1

incomplete

7/17/14: Transfect MAV1212-hEF1a-eBFP2 (1084.0 ng/uL), Mutant hEF1a-2xKturn-eGFP (551.4 ng/uL) and Wild Type hEF1a-2xKturn-eGFP (287.0 ng/uL), MAV1212-hEF1a-L7ae (1686.4 ng/uL) into HEK cells

7

incomplete

7/19/14 (2 pm): Flow Cytometer

DNA Dilution: 5 uL MAV1212-hEF1a-eBFP2, 10 uL Mutant Hef1a-2xKturn-eGFP, 20 uL Wild Type Hef1a-2xKturn-eGFP, 3 uL MAV1212-hEF1a-L7ae

24-WELL PLATE TRANSFECTION MAP:

(B)1

MAV1212-hEF1a-eBFP2

(B)₂

MAV1212-hEF1a-eBFP2

//////

(G_M)1

Mutant Hef1a-2xKturn-eGFP

(G_M)₂

Mutant Hef1a-2xKturn-eGFP

(G_M+ B)₁

Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_M+ B)₂

Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_M+ B + L)₁

Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2
hEF1a-L7ae

(G_M+ B + L)₂

Mutant Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2
hEFa-L7ae

(G_WT)1

Wild Type Hef1a-2xKturn-eGFP

(G_WT)₂

Wild Type Hef1a-2xKturn-eGFP

(G_WT+ B)₁

Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_WT+ B)₂

Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2

(G_WT+ B + L)₁

Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2
hEFa1-L7ae

(G_WT+ B + L)₂

Wild Type Hef1a-2xKturn-eGFP
MAV1212-hEF1a-eBFP2
hEF1a-L7ae

Neg Ctrl

Dummy DNA
With Lipofectamine

Neg Ctrl

Dummy DNA
With Lipofectamine

Neg/Neg Ctrl

No DNA
No Lipofectamin

Neg/Neg Ctrl

No DNA
No Lipofectamine

//////

RESULT: Brief description of experimental outcome (mean transfection efficiency and implications). Link to results page with data analysi

Experiment #2

GOAL: Test the strength of TAL14

14

incomplete

Day 1: Seed cells

15

incomplete

Day 2: Transfect MAV1212-hEF1a-eBFP2 (1084.0 ng/uL), UAS-TAL14-mKate (___ ng/uL), hEF1a-GAL4VP16 (___ ng/uL), hEF1a-TAL14 (___ ng/uL) and UAS-TAL14-eYFP (___ ng/uL) into HEK cells

16

incomplete

Day 34: Flow Cytometer

DNA Dilution: 5 uL MAV1212-hEF1a-eBFP2, ___ uL UAS-TAL14-mKate, ___ uL hEF1a-GAL4VP16, ___ uL hEF1a-TAL14, ___ UAS-TAL14-eYFP

24-WELL PLATE TRANSFECTION MAP:

(B)₁

- MAV1212-hEF1a-eBFP2

(B)₂

- MAV1212-hEF1a-eBFP2

//////

Neg Ctrl

- Dummy DNA
- With Lipofectamine

(B + Y)₁

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-eYFP

(B + Y)₂

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-eYFP

(B + R)₁

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-mKate

(B + R)₂

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-mKate

//////

Neg Ctrl

- Dummy DNA
- With Lipofectamine

(B + Y + GAL)₁

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-eYFP
- hEF1a-Gal4VP16

(B + Y + GAL)₂

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-eYFP
- hEF1a-Gal4VP16

(B + R + GAL)₁

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-mKate
- hEF1a-Gal4VP16

(B + R + GAL)₂

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-mKate
- hEF1a-Gal4VP16

//////

Neg/Neg Ctrl

- No DNA
- No Lipofectamin

(B + Y + GAL)₁

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-eYFP
- hEF1a-Gal4VP16
- hEF1a-TAL14

(B + Y + GAL)₂

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-eYFP
- hEF1a-Gal4VP16
- hEF1a-TAL14

(B + R + GAL)₁

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-mKate
- hEF1a-Gal4VP16
- hEF1a-TAL14

(B + R + GAL)₂

- MAV1212-hEF1a-eBFP2
- UAS-TAL14-mKate
- hEF1a-Gal4VP16
- hEF1a-TAL14

//////

Neg/Neg Ctrl

- No DNA
- No Lipofectamine

RESULT: Brief description of experimental outcome (mean transfection efficiency and implications). Link to results page with data analysis.

Experiment #3

GOAL: Test the sensors in absence of siRNA

10

incomplete

Day 1: Seed cells

11

incomplete

Day 2: Transfect Low Sensor miR-144 (___ ng/uL), Low Sensor miR-181c (___ ng/uL), High Sensor miR-30d (___ ng/uL), High Sensor miR-146a (___ ng/uL), High Sensor 125b (___ ng/uL) and High Sensor let-7f (___ ng/uL) into HEK cells

12

incomplete

Day 4: (2 pm): Flow Cytometer

DNA Dilution: ___ uL Low Sensor miR-144, ___ uL Low Sensor miR-181c, ___ uL High Sensor miR-30d, ___ uL High Sensor miR-146a, ___ uL High Sensor 125b, ___ uL High Sensor let-7f

24-WELL PLATE TRANSFECTION MAP:

144

- Low Sensor miR-144

- Low Sensor miR-181c

- High Sensor miR-30d

- High Sensor miR-146a

- High Sensor miR-125b

- High Sensor let-7f

- Low Sensor miR-144

- Low Sensor miR-181c

- High Sensor miR-30d

- High Sensor miR-146a

- High Sensor miR-125b

- High Sensor let-7f

//////

Neg Ctrl

- Dummy DNA
- With Lipofectamine

Neg Ctrl

- Dummy DNA
- With Lipofectamine

Neg/Neg Ctrl

- No DNA
- No Lipofectamine

Neg/Neg Ctrl

- No DNA
- No Lipofectamine

//////

RESULT: Brief description of experimental outcome (mean transfection efficiency and implications). Link to results page with data analysis.

Experiment #4

GOAL: Test the sensors in the presence of siRNA

13

incomplete

Day 1: Seed cells

9

incomplete

Day 2: Transfect Low Sensor miR-144 (___ ng/uL), Low Sensor miR-181c (___ ng/uL), High Sensor miR-30d (___ ng/uL), High Sensor miR-146a (___ ng/uL), High Sensor 125b (___ ng/uL), High Sensor let-7f (___ ng/uL), Mature 144 (___ ng/uL), Mature 181c (___ ng/uL), Mature 30d (___ ng/uL), Mature 146a (___ ng/uL), Mature 125b (___ ng/uL) and Mature let-7f (___ ng/uL) into HEK cells

9

incomplete

Day 4: (2 pm): Flow Cytometer

DNA Dilution: ___ uL Low Sensor miR-144, ___ uL Low Sensor miR-181c, ___ uL High Sensor miR-30d, ___ uL High Sensor miR-146a, ___ uL High Sensor 125b, ___ uL High Sensor let-7f , ___ uL Mature 144, ___ uL Mature 181c, ___ uL Mature 30d, ___ uL Mature 146a, ___ uL Mature 125b and ___ uL Mature let-7f

24-WELL PLATE TRANSFECTION MAP:

144

- Low Sensor miR-144
- Mature 144

- Low Sensor miR-181c
- Mature 181c

- High Sensor miR-30d
- Mature 30d

- High Sensor miR-146a
- Mature 146a

- High Sensor miR-125b
- Mature 125b

- High Sensor let-7f
- Mature let-7f

- Low Sensor miR-144
- Mature 144

- Low Sensor miR-181c
- Mature 181c

- High Sensor miR-30d
- Mature 30d

- High Sensor miR-146a
- Mature 146a

- High Sensor miR-125b
- Mature 125b

- High Sensor let-7f
- Mature let-7f

//////

Neg Ctrl

- Dummy DNA
- With Lipofectamine

Neg Ctrl

- Dummy DNA
- With Lipofectamine

Neg/Neg Ctrl

- No DNA
- No Lipofectamine

Neg/Neg Ctrl

- No DNA
- No Lipofectamine

//////

RESULT: Brief description of experimental outcome (mean transfection efficiency and implications). Link to results page with data analysis.

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