Team:IvyTech-South Bend/Notebook

From 2010.igem.org

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(9/16/10)
(9/17/10)
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Today we will be doing a DNA Extraction from/of part T9002 for electroporation this will determine if we have a stable host for our beast
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Today we will be doing a DNA Extraction from/of part T9002 for electroporation this will determine if we have a stable host for our beast this will also Work!
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this will also Work!
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Following protocols from pgs 15 &19
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-instead of Electroporating our part we will use PG10 to determine if these lines are good hosts
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First we will be pouring a full pack of LB/Agar/Amp plates using the LB/Agar I made on 9/2/10 then adding amp made by DG on 9/1/10 we will also add 5 mL Arabonse to the Agar before pouring plates have been poured now letting stand for an hour to harden.
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-Now I will be making 1L of LB/Broth for Culturing.
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Lot 9082989 ex. 2014-02-28
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BD Difco LB/Broth – Lennox 20 g per 1L
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so I will put 10 g into 2 elenmyier flask/capped one weighed 10.0023 g and the other weighed 10.0011 g then I added 500 mL into each flask then shook to mix then dissolved in microwaved for 8 oz setting once then placed into Autoclave for 15 min @ 121 degrees C until done.
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Now we are going electroporate our grown t/pos Electro Comp Cells from yesterday with PG10 using protocol from pg 15 changing # 5 to 5 mL of PG10
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We will be using a different protocol for electroporation of grown t/pos bacteria from Bio-Rad’s website prof. T will print and I’ll place here.
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The mycobacterium will be done by electroporation protocol on pg 15.
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1) First I placed a Cuvette into Ice bath and thawed electro comp cells
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2) I set up 5 1.5 mL Sterile Centrifuge tubes  with 900 mL SOB media
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3) we will electroporate Irene’s electrocomp cells w/PG10 and mine will contain the parts after electroporation.
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Code
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AT – A.tumefacions SL – S.lactis BM – B.magetanium MP – Mycobacteria
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BT – B.thurigensis
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split cells using 40 mL Hy part my cells then electroporate
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I electroporated all except mycobacteria now letting cells sit for at least
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Just added 1 mL to electroporated Cultures they had sat for 20 min before adding.
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- Now I will electroporate mycobacteria using E – Coli protocol from pg 15 using 40 mL cells 4 mL pg10 Electroporation hit twice Read out 1.8 kv @ 5.2 ms then 1.79 kv @ 5.1 ms. then placed into 900 mL SOB media then incubate @ 30 degrees C for 3 hrs.
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Results
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From this we found that agrobacteria will be the best host for our beast/IGem
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we will do further testing towards our ultimate goal.
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== 9/21/10 ==
== 9/21/10 ==

Revision as of 14:09, 12 October 2010

Discussion

Contents

Notebook

June
MTWTFSS
  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
July
MTWTFSS
      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
August
MTWTFSS
            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



September
MTWTFSS
    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
October
MTWTFSS
        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
November
MTWTFSS
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

8/24/10

Today we will be pouring new plates for streaking new transformed cells.

8/27/10

We will be running a gel to determine if our DNA sample was successfully electroplated.

8/27/10

Today we will be electroplating part KBBa_3131010

8/31/10

Results of transformation - the plates had growth but (no) distinct colony pattern.

8/31/10

After agar is dissolved, and allowed to cool, I will add Amp in different doses to specific plates. JH

9/1/10

Today the team will be making up the same plates as I made yesterday. Professor T. wants new Amp stock made to document how much is used into the stock.

9/2/10

Results from streaking –

9/2/10

Protocol For Making LB-Broth/Agar


9/3/10

Today I’m finishing making the LB/Agar from yesterday.

9/9/10

Due to conflict we will be changing from E Coli to yeast.

9/10/10

Lux Casette Right Promoter BBa_I1051

9/14/10

Today we found growth from our Electroporated E – Coli parts

9/15/10

- Today we will be extracting DNA from out electroporated cells/T9002


9/16/10

Today we will wash

9/17/10

Today we will be doing a DNA Extraction from/of part T9002 for electroporation this will determine if we have a stable host for our beast this will also Work!

9/21/10

Today we will electroporate Agrobacterium with part T9002 using protocol from pg 22 – 23

9/22/10

Today we have growth from electroporation done yesterday we are going to test by hitting with (AHL) the 20 mL grew not 5 mL sample

9/22/10

The only one that should glow should be (C)/(3) E – Coli w/transformed part in LB For Supernate of A.Tumafaciens

9/23/10

Today I have been setting up for the trial of our part T9002 inside agrobacteria to see if in the presence of AHL it floureces.

9/24/10

Today I’m testing absorbance of our trials @ 395nm

9/28/10

Today we will be picking 2 colonies from the Lacz, LuxR electroporated plates that were streaked on 9/22/10



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