Sensor: Week 2 May 23-27

From 2011.igem.org

(Difference between revisions)

Latest revision as of 15:35, 10 June 2011

Monday

cI Repressor Test Assembly, Day 1

The sensor group began the assembly of a cI repressor testing construct by amplifying the below constitutive promoters via PCR.

Assembly Step	Parts Involved in Assembly Step
PCR	J23113 J23105 J23118 J23100

Tuesday

cI Repressor Test Assembly, Day 2

Jamie says: "No more research. To the lab!"

The sensor group continued to build their test construct by inserting the constitutive promoters into the vector containing the cI repressor and ribosome binding site (B0034+C0051). The sensor group also inserted a ribosome binding site and color reporter (B0034+mCherry) in front of a terminator (B0015).

Assembly Step	Parts Involved in Assembly Step
PCR Purification	J23113 J23105 J23118 J23100
Miniprep	B0051
Restriction Digest	Inserts using EcoRI and SpeI:	J23113 J23105 J23118 J23100 B0034+mCherry
	Vectors using EcoRI and XbaI:	B0034+C0051 B0015
Ligation	J23113 and B0034+C0051 J23105 and B0034+C0051 J23118 and B0034+C0051 J23100 and B0034+C0051 B0034+mCherry and B0015
Transformation/Plating	The above ligations were transformed into Escherichia coli cells and plated onto Amp resistant plates.
PCR	B0030 B0031 B0032 B0033

Wednesday

cI Repressor Test Assembly, Day 3

Jamie says: "D'oh! No one said anything about WAITING!"

The sensor group inserted various ribosome binding sites (listed below) in front of the cI repressor (C0051). The sensor group also finished the assemblies from yesterday by performing colony PCR on the plated colonies.

Assembly Step	Parts Involved with Assembly Step
Colony PCR	B0034+mCherry+B0015 (Kan resistance) B0034+mCherry+B0015 (Amp resistance)
PCR Purification	B0030 B0031 B0032 B0033
Restriction Digest	Inserts using EcoRI and SpeI:	B0030 B0031 B0032 B0033
Restriction Digest	Vector using EcoRI and XbaI:	C0051
Ligation	B0030+C0051 B0031+C0051 B0032+C0051 B0033+C0051
Transformation/Plating	The above ligations were transformed into Escherichia coli cells and plated onto Amp resistant plates.

Thursday

cI Repressor Test Assembly, Day 4

Finally, the oligos arrived!

The sensor group put the reporting assembly (B0034+mCherry+B0015) in front of two cI repressor construct with different constitutive promoters (J23113+B0034+C0051 and J23105+B0034+C0051).

Assembly Step	Part Involved with Assembly Step
Miniprep for Sequencing	B0034+mCherry+B0015 J23113+B0034+C0051 J23105+B0034+C0051
PCR	B0034+mCherry+B0015
Restriction Digest	Insert using EcoRI and SpeI:	B0034+mCherry+B0015
Restriction Digest	Vectors using EcoRI and XbaI:	J23113+B0034+C0051 J23105+B0034+C0051
Ligation	B0034+mCherry+B0015 and J23113+B0034+C0051 B0034+mCherry+B0015 and J23105+B0034+C0051
Transformation/Plating	The above ligations were transformed into Escherichia coli cells and plated onto Amp resistant plates.

Note: The minipreps in green font indicate the sequencing proved the assembly worked.

Friday

cI Repressor Test Assembly, Day 5

The week's assemblies have been confirmed and named for the registry.

Assembly	Registry Name
B0034+mCherry+B0015 J23113+B0034+C0051 J23105+B0034+C0051	K648000 K648001 K648002

Back to Notebook

@@ Line 1: / Line 1: @@
 ==Monday==
-===Constitutive Promoters for Testing cI Repressor, Day 1===
+===cI Repressor Test Assembly, Day 1===
+The sensor group began the assembly of a cI repressor testing construct by amplifying the below constitutive promoters via PCR.
 {|border="1"
 !Assembly Step
@@ Line 10: / Line 11: @@
 ==Tuesday==
+===cI Repressor Test Assembly, Day 2===
 [[File:Jamie_research.jpg|thumb|right|400px|Jamie says: "No more research. To the lab!"]]
+The sensor group continued to build their test construct by inserting the constitutive promoters into the vector containing the cI repressor and ribosome binding site (B0034+C0051). The sensor group also inserted a ribosome binding site and color reporter (B0034+mCherry) in front of a terminator (B0015).
 {|border="1"
 |'''Assembly Step'''
@@ Line 23: / Line 26: @@
 |rowspan="2" | Restriction Digest
 |Inserts using EcoRI and SpeI:
-|J23113<br />J23105<br />J23118<br />J23100<br />B0034+Mcherry
+|J23113<br />J23105<br />J23118<br />J23100<br />B0034+mCherry
 |-
 |Vectors using EcoRI and XbaI:
@@ Line 29: / Line 32: @@
 |-
 |Ligation
-|colspan="2" align="center" | J23113 and B0034+C0051<br />J23105 and B0034+C0051<br />J23118 and B0034+C0051<br />J23100 and B0034+C0051<br />B0034+Mcherry and B0015
+|colspan="2" align="center" | J23113 and B0034+C0051<br />J23105 and B0034+C0051<br />J23118 and B0034+C0051<br />J23100 and B0034+C0051<br />B0034+mCherry and B0015
 |-
 |Transformation/Plating
@@ Line 39: / Line 42: @@
 ==Wednesday==
+===cI Repressor Test Assembly, Day 3===
 [[File:Everyone_lab_waiting_safe.jpg|right|thumb|400px|Jamie says: "D'oh! No one said anything about WAITING!"]]
+The sensor group inserted various ribosome binding sites (listed below) in front of the cI repressor (C0051). The sensor group also finished the assemblies from yesterday by performing colony PCR on the plated colonies.
 {|border="1"
 !Assembly Step
@@ Line 45: / Line 50: @@
 |-
 |Colony PCR
-|colspan="2" | B0034+Mcherry+B0015 (Kan resistance)<br />B0034+Mcherry+B0015 (Amp resistance)
+|colspan="2" | B0034+mCherry+B0015 (Kan resistance)<br />B0034+mCherry+B0015 (Amp resistance)
 |-
 |PCR Purification
@@ Line 65: / Line 70: @@
 ==Thursday==
+===cI Repressor Test Assembly, Day 4===
 [[File:Oligos_arrival.jpg|thumb|right|350px|Finally, the oligos arrived!]]
+The sensor group put the reporting assembly (B0034+mCherry+B0015) in front of two cI repressor construct with different constitutive promoters (J23113+B0034+C0051 and J23105+B0034+C0051).
 {|border="1"
 !Assembly Step
@@ Line 92: / Line 99: @@
 ==Friday==
-The following assemblies have been confirmed and named for the registry.
+===cI Repressor Test Assembly, Day 5===
+The week's assemblies have been confirmed and named for the registry.
 {|border="1"
 !Assembly