Template:Team:UC Berkeley/Notebook/MJV construction

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 +
[[Template:Team:UC Berkeley/Notebook/MJV construction/corrected assembly vector | patching attR2 site in assembly vector]]<br>
 +
[[Template:Team:UC Berkeley/Notebook/MJV construction/OriV entry vector | OriV entry vector]]
 +
 +
'''Construction of {<Cre!} Basic Part K112100'''
'''Construction of {<Cre!} Basic Part K112100'''
<pre>
<pre>
-
PCR mv001/mv002 on pBca9145-Bca1117   (1060bp, EcoRI/BamHI/DpnI)
+
PCR mv001/mv002 on pBca9145-Bca1010   (1060bp, gp, EcoRI/BamHI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112100
Product is pBca1256-K112100
-----------------------------------------------
-----------------------------------------------
mv001 Forward Biobricking of <Cre!
mv001 Forward Biobricking of <Cre!
-
ccataGAATTCatgAGATCTtccaatttactgaccgtacac
+
ccataGAATTCatgAGATCTtccaatttactgaccgtacacc
mv002 Reverse Biobricking of <Cre!
mv002 Reverse Biobricking of <Cre!
-
cgttaGGATCCttaatcgccatcttccagcaggcg
+
cgttaGGATCCttaatcgccatcttccagc
</pre>
</pre>
'''Construction of {Cre>} Basic Part K112101'''
'''Construction of {Cre>} Basic Part K112101'''
<pre>
<pre>
-
PCR mv003/mv004 on pBca9145-Bca1117   (1060bp, EcoRI/BamHI/DpnI)
+
PCR mv003/mv004 on pBca9145-Bca1010   (1060bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112101
Product is pBca1256-K112101
Line 26: Line 30:
'''Construction of {<Cre>} Basic Part K112102'''
'''Construction of {<Cre>} Basic Part K112102'''
<pre>
<pre>
-
PCR mv001/mv004 on pBca9145-Bca1117   (1057bp, EcoRI/BamHI/DpnI)
+
PCR mv001/mv004 on pBca9145-Bca1010   (1057bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112102
Product is pBca1256-K112102
-----------------------------------------------
-----------------------------------------------
mv001 Forward Biobricking of <Cre!
mv001 Forward Biobricking of <Cre!
-
ccataGAATTCatgAGATCTtccaatttactgaccgtacac
+
ccataGAATTCatgAGATCTtccaatttactgaccgtacacc
mv004 Reverse Biobricking of Cre>
mv004 Reverse Biobricking of Cre>
cgttaGGATCCatcgccatcttccagcaggcg
cgttaGGATCCatcgccatcttccagcaggcg
Line 37: Line 41:
-
'''Construction of {r.BamH1!} Basic Part K112103'''
+
'''Construction of {r.BamHI!} Basic Part K112103'''
<pre>
<pre>
PCR mv005/mv006 on pBca9145-I716210    (673bp, EcoRI/BamHI/DpnI)
PCR mv005/mv006 on pBca9145-I716210    (673bp, EcoRI/BamHI/DpnI)
Line 44: Line 48:
-----------------------------------------------
-----------------------------------------------
mv005 Forward Biobricking of r.BamH1!
mv005 Forward Biobricking of r.BamH1!
-
ccataGAATTCatgAGATCTatggaagtagaaaaagag
+
ccataGAATTCatgAGATCTatggaagtagaaaaagagtttattactgatgaagc
mv006 Reverse Biobricking of r.BamH1!
mv006 Reverse Biobricking of r.BamH1!
-
cgttaGGATCCctatttgttttcaactttatc
+
cgttaGGATCCctatttgttttcaactttatctttcc
</pre>
</pre>
Line 56: Line 60:
-----------------------------------------------
-----------------------------------------------
mv005 Forward Biobricking of r.BamHI!
mv005 Forward Biobricking of r.BamHI!
-
ccataGAATTCatgAGATCTatggaagtagaaaaagag
+
ccataGAATTCatgAGATCTatggaagtagaaaaagagtttattactgatgaagc
mv007 Reverse Biobricking of r.BamHI>
mv007 Reverse Biobricking of r.BamHI>
cgttaGGATCCtttgttttcaactttatctttcc
cgttaGGATCCtttgttttcaactttatctttcc
Line 68: Line 72:
-----------------------------------------------
-----------------------------------------------
mv008 Forward Biobricking of <r.BamHI>
mv008 Forward Biobricking of <r.BamHI>
-
ccataGAATTCatgAGATCTgaagtagaaaaagagtttattac
+
ccataGAATTCatgAGATCTgaagtagaaaaagagtttattactgatg
mv007 Reverse Biobricking of r.BamHI>
mv007 Reverse Biobricking of r.BamHI>
cgttaGGATCCtttgttttcaactttatctttcc
cgttaGGATCCtttgttttcaactttatctttcc
Line 80: Line 84:
-----------------------------------------------
-----------------------------------------------
mv009 Forward Biobricking of r.BglII!
mv009 Forward Biobricking of r.BglII!
-
ccataGAATTCatgAGATCTatgaagattgatataacggactataacc
+
ccataGAATTCatgAGATCTatgaagattgatataacggactataaccatgc
mv010 Reverse Biobricking of r.BglII!
mv010 Reverse Biobricking of r.BglII!
cgttaGGATCCttaatatgtcacgattgttcctc
cgttaGGATCCttaatatgtcacgattgttcctc
Line 92: Line 96:
-----------------------------------------------
-----------------------------------------------
mv009 Forward Biobricking of r.BglII!
mv009 Forward Biobricking of r.BglII!
-
ccataGAATTCatgAGATCTatgaagattgatataacggactataacc
+
ccataGAATTCatgAGATCTatgaagattgatataacggactataaccatgc
mv011 Reverse Biobricking of r.BglII>
mv011 Reverse Biobricking of r.BglII>
cgttaGGATCCatatgtcacgattgttcctc
cgttaGGATCCatatgtcacgattgttcctc
Line 104: Line 108:
-----------------------------------------------
-----------------------------------------------
mv012 Forward Biobricking of <r.BglII>
mv012 Forward Biobricking of <r.BglII>
-
ccataGAATTCatgAGATCTaagattgatataacggac
+
ccataGAATTCatgAGATCTaagattgatataacggactataacc
mv011 Reverse Biobricking of r.BglII>
mv011 Reverse Biobricking of r.BglII>
cgttaGGATCCatatgtcacgattgttcctc
cgttaGGATCCatatgtcacgattgttcctc
Line 114: Line 118:
PCR mv015/mv018 on T4 DNA ligase    (376 bp, gp = B)
PCR mv015/mv018 on T4 DNA ligase    (376 bp, gp = B)
PCR mv017/mv020 on T4 DNA ligase    (191 bp, gp = C)
PCR mv017/mv020 on T4 DNA ligase    (191 bp, gp = C)
-
PCR mvo19/mv014 on T4 DNA ligase    (746 bp, gp = D)
+
PCR mv019/mv014 on T4 DNA ligase    (746 bp, gp = D)
---------------------------------------------------
---------------------------------------------------
PCR mv013/mv018 on A+B              (622 bp, gp = E)
PCR mv013/mv018 on A+B              (622 bp, gp = E)
Line 124: Line 128:
-----------------------------------------------
-----------------------------------------------
mv013 Forward Biobricking of rbs.ligase!
mv013 Forward Biobricking of rbs.ligase!
-
ccataGAATTCatgAGATCTaaactctaaaggatgaac
+
ccataGAATTCatgAGATCTaaactctaaaggatgaacaatg
-
mv014 Reverse Biobricking of rbs.ligase!
+
mv014 Reverse Biobricking of ligase!
cgttaGGATCCtcatagaccagttacctcatg
cgttaGGATCCtcatagaccagttacctcatg
mv015  Removing the 1st EcoRI site in T4 DNA ligase(F)  gacttcattgaGttcacattagc
mv015  Removing the 1st EcoRI site in T4 DNA ligase(F)  gacttcattgaGttcacattagc
Line 134: Line 138:
mv020  Removing the 2nd EcoRI site in T4 DNA ligase(R)  gctacttcggcgaaCtctttagc
mv020  Removing the 2nd EcoRI site in T4 DNA ligase(R)  gctacttcggcgaaCtctttagc
</pre>
</pre>
 +
 +
 +
'''Construction of {ligase!} Basic Part K112110'''
 +
<pre>
 +
PCR mv021/mv014 on pBca1256-K112109    (1495bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112110
 +
-----------------------------------------------
 +
mv021 Forward Biobricking of ligase!
 +
ccataGAATTCatgAGATCTatgattcttaaaattctgaacg
 +
mv014 Reverse Biobricking of ligase!
 +
cgttaGGATCCtcatagaccagttacctcatg
 +
</pre>
 +
 +
'''Construction of {<ligase!} Basic Part K112111'''
 +
<pre>
 +
PCR mv022/mv014 on pBca1256-K112109    (1492bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112111
 +
-----------------------------------------------
 +
mv022 Forward Biobricking of <ligase!
 +
ccataGAATTCatgAGATCTattcttaaaattctgaacg
 +
mv014 Reverse Biobricking of ligase!
 +
cgttaGGATCCtcatagaccagttacctcatg
 +
</pre>
 +
 +
'''Construction of {ligase>} Basic Part K112112'''
 +
<pre>
 +
PCR mv021/mv023 on pBca1256-K112109    (1492bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112112
 +
-----------------------------------------------
 +
mv021 Forward Biobricking of ligase!
 +
ccataGAATTCatgAGATCTatgattcttaaaattctgaacg
 +
mv023 Reverse Biobricking of ligase>
 +
cgttaGGATCCtagaccagttacctcatg
 +
</pre>
 +
 +
'''Construction of {<ligase>} Basic Part K112113'''
 +
<pre>
 +
PCR mv022/mv023 on pBca1256-K112109    (1489bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112113
 +
-----------------------------------------------
 +
mv022 Forward Biobricking of <ligase!
 +
ccataGAATTCatgAGATCTattcttaaaattctgaacg
 +
mv023 Reverse Biobricking of ligase>
 +
cgttaGGATCCtagaccagttacctcatg
 +
</pre>
 +
 +
'''Construction of {a~ligase!} Basic Part K112114'''
 +
<pre>
 +
PCR mv024/mv014 on pBca1256-K112109    (1499bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112114
 +
-----------------------------------------------
 +
mv024 Forward Biobricking of a~ligase!
 +
ccataGAATTCatgAGATCTaacaatgattcttaaaattctgaacg
 +
mv014 Reverse Biobricking of ligase!
 +
cgttaGGATCCtcatagaccagttacctcatg
 +
</pre>
 +
 +
'''Construction of {a~Cre!} Basic Part K112115'''
 +
<pre>
 +
PCR mv025/mv002 on pBca9145-Bca1010    (1067bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112115
 +
-----------------------------------------------
 +
mv025 Forward Biobricking of a~Cre!
 +
ccataGAATTCatgAGATCTttaaatgtccaatttactgaccgtacacc
 +
mv002 Reverse Biobricking of <Cre!
 +
cgttaGGATCCttaatcgccatcttccagc
 +
</pre>
 +
 +
'''Construction of {a~r.BamHI!} Basic Part K112116'''
 +
<pre>
 +
PCR mv026/mv006 on pBca9145-I716210    (677bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112116
 +
-----------------------------------------------
 +
mv026 Forward Biobricking of a~r.BamH1!
 +
ccataGAATTCatgAGATCTtagaatggaagtagaaaaagagtttattactgatgaagc
 +
mv006 Reverse Biobricking of r.BamH1!
 +
cgttaGGATCCctatttgttttcaactttatctttcc
 +
</pre>
 +
 +
'''Construction of {a~r.BglII!} Basic Part K112117'''
 +
<pre>
 +
PCR mv027/mv010 on pBca9145-Bca1147    (707bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112117
 +
-----------------------------------------------
 +
mv027 Forward Biobricking of a~r.BglII!
 +
ccataGAATTCatgAGATCTcataatgaagattgatataacggactataaccatgc
 +
mv010 Reverse Biobricking of r.BglII!
 +
cgttaGGATCCttaatatgtcacgattgttcctc
 +
</pre>
 +
 +
'''Construction of rrnB P1 promoter Basic Part K112118'''
 +
<pre>
 +
PCR mv028/mv029 on rrnB native sequence  (721bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                        (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112118
 +
-----------------------------------------------
 +
mv028 Forward Biobricking of rrnB P1 promoter
 +
cgataGAATTCatgAGATCTatgcactaaaacgtatcctacgcccg
 +
mv029  Reverse Biobricking of rrnB P1 promoter
 +
cgttaGGATCCcgccttcccgctacagagtcaagc
 +
</pre>
 +
 +
'''Construction of {rbs.Cre!} Basic Part K112119'''
 +
<pre>
 +
PCR mv030/mv002 on pBca9145-Bca1010    (1082bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112119
 +
-----------------------------------------------
 +
mv030 Forward Biobricking of rbs.Cre!
 +
cgataGAATTCatgAGATCTaccattatctgagtgttaaatgtccaatttactgaccgtacacc
 +
mv002 Reverse Biobricking of <Cre!
 +
cgttaGGATCCttaatcgccatcttccagc
 +
</pre>
 +
 +
 +
'''Construction of {rbs.r.BamHI!} Basic Part K112120'''
 +
<pre>
 +
PCR mv031/mv006 on pBca9145-I716210    (692bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112120
 +
-----------------------------------------------
 +
mv031 Forward Biobricking of rbs.r.BamH1!
 +
cgataGAATTCatgAGATCTtaagtgggggcaaattagaatggaagtagaaaaagagtttattactgatgaagc
 +
mv006 Reverse Biobricking of r.BamH1!
 +
cgttaGGATCCctatttgttttcaactttatctttcc
 +
</pre>
 +
 +
'''Construction of {rbs.r.BglII!} Basic Part K112121'''
 +
<pre>
 +
PCR mv032/mv010 on pBca9145-Bca1147    (722bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112121
 +
-----------------------------------------------
 +
mv032 Forward Biobricking of rbs.r.BglII!
 +
cgataGAATTCatgAGATCTtggaaggagacactcataatgaagattgatataacggactataaccatgc
 +
mv010 Reverse Biobricking of r.BglII!
 +
cgttaGGATCCttaatatgtcacgattgttcctc
 +
</pre>
 +
 +
'''Construction of {Cre!} Basic Part K112122'''
 +
<pre>
 +
PCR mv003/mv002 on pBca9145-Bca1010    (1063bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112122
 +
-----------------------------------------------
 +
mv003 Forward Biobricking of Cre>
 +
ccataGAATTCatgAGATCTatgtccaatttactgaccgtac
 +
mv002 Reverse Biobricking of <Cre!
 +
cgttaGGATCCttaatcgccatcttccagc
 +
</pre>
 +
 +
'''Construction of {<r.BglII!} Basic Part K112123'''
 +
<pre>
 +
PCR mv012/mv010 on pBca9145-Bca1147    (700bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112123
 +
-----------------------------------------------
 +
mv012 Forward Biobricking of <r.BglII>
 +
ccataGAATTCatgAGATCTaagattgatataacggactataacc
 +
mv010 Reverse Biobricking of r.BglII!
 +
cgttaGGATCCttaatatgtcacgattgttcctc
 +
</pre>
 +
 +
'''Construction of {rbs.trfA!} Basic Part K112124'''
 +
<pre>
 +
PCR JH102F/JH102R on RP4 genome      (416bp, EcoRI/BamHI/DpnI)
 +
Sub into pBjh1601KC                  (EcoRI/BamHI, 3132+778, L)
 +
Product is pBjh1601KC-K112124
 +
-----------------------------------------------
 +
JH102F Forward Biobricking of {rbs.trfA!}
 +
ccatgGAATTCatgAGATCTgcgaggggcggcttccgctAtgtacaaccag
 +
JH102R Reverse Biobricking of {rbs.trfA!}
 +
cgttaGGATCCtcagagccttccacgcagc
 +
</pre>
 +
 +
'''Construction of {OriV} Basic Part K112125'''
 +
<pre>
 +
PCR mv037/mv038 on pSMART Fosmid Blunt Vector (Lucigen)    (677bp, EcoRI/BamHI/DpnI)
 +
Sub into pBca1256                        (EcoRI/BamHI, 2472+910, L)
 +
Product is pBca1256-K112125
 +
-----------------------------------------------
 +
mv037  Forward biobricking of OriV from pSMART
 +
cgataGAATTCatgAGATCTtgcttcctcgctcactcgac
 +
mv038  Reverse biobricking of OriV form pSMART
 +
gctagGGATCCaagccctcggctggttgc
 +
</pre>
 +
 +
'''Construction of {temp. sensitive promoter} Basic Part K112126'''
 +
<pre>
 +
PCR JH115F/JH115R on pAH57
 +
sub into pBca1256
 +
Product is pBca1256-K112126
 +
-----------------------------------------------
 +
JH115F  Forward biobricking of cI, Pr
 +
cgataGAATTCatgAGATCTgaagattcttgctcaattgttatcagc
 +
JH115R  Reverse biobricking of cI, Pr
 +
gctagGGATCCagaattgatacaacctccttagtacatgc
 +
</pre>
 +
 +
----
 +
 +
<div style="text-align: center;"> [[Team:UC_Berkeley/Notebook/Madhvi_Venkatesh|Madhvi's Notebook]] </div>
 +
<div style="text-align: center;"> [[Team:UC_Berkeley|UC Berkeley Team Homepage]] </div>

Latest revision as of 23:25, 22 September 2008

patching attR2 site in assembly vector
OriV entry vector


Construction of {<Cre!} Basic Part K112100 

PCR mv001/mv002 on pBca9145-Bca1010    (1060bp, gp, EcoRI/BamHI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112100
-----------------------------------------------
mv001	Forward Biobricking of <Cre!
ccataGAATTCatgAGATCTtccaatttactgaccgtacacc
mv002	Reverse Biobricking of <Cre!
cgttaGGATCCttaatcgccatcttccagc

Construction of {Cre>} Basic Part K112101 

PCR mv003/mv004 on pBca9145-Bca1010    (1060bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112101
-----------------------------------------------
mv003	Forward Biobricking of Cre>
ccataGAATTCatgAGATCTatgtccaatttactgaccgtac
mv004	Reverse Biobricking of Cre>
cgttaGGATCCatcgccatcttccagcaggcg


Construction of {<Cre>} Basic Part K112102 

PCR mv001/mv004 on pBca9145-Bca1010    (1057bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112102
-----------------------------------------------
mv001	Forward Biobricking of <Cre!
ccataGAATTCatgAGATCTtccaatttactgaccgtacacc
mv004	Reverse Biobricking of Cre>
cgttaGGATCCatcgccatcttccagcaggcg


Construction of {r.BamHI!} Basic Part K112103 

PCR mv005/mv006 on pBca9145-I716210    (673bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112103
-----------------------------------------------
mv005	Forward Biobricking of r.BamH1!
ccataGAATTCatgAGATCTatggaagtagaaaaagagtttattactgatgaagc
mv006	Reverse Biobricking of r.BamH1!
cgttaGGATCCctatttgttttcaactttatctttcc

Construction of {r.BamHI>} Basic Part K112104 

PCR mv005/mv007 on pBca9145-I716210    (670bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112104
-----------------------------------------------
mv005	Forward Biobricking of r.BamHI!
ccataGAATTCatgAGATCTatggaagtagaaaaagagtttattactgatgaagc
mv007	Reverse Biobricking of r.BamHI>
cgttaGGATCCtttgttttcaactttatctttcc

Construction of {<r.BamHI>} Basic Part K112105 

PCR mv008/mv007 on pBca9145-I716210    (667bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112105
-----------------------------------------------
mv008	Forward Biobricking of <r.BamHI>
ccataGAATTCatgAGATCTgaagtagaaaaagagtttattactgatg
mv007	Reverse Biobricking of r.BamHI>
cgttaGGATCCtttgttttcaactttatctttcc

Construction of {r.BglII!} Basic Part K112106 

PCR mv009/mv010 on pBca9145-Bca1147    (703bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112106
-----------------------------------------------
mv009	Forward Biobricking of r.BglII!
ccataGAATTCatgAGATCTatgaagattgatataacggactataaccatgc
mv010	Reverse Biobricking of r.BglII!
cgttaGGATCCttaatatgtcacgattgttcctc

Construction of {r.BglII>} Basic Part K112107 

PCR mv009/mv011 on pBca9145-Bca1147    (700bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112107
-----------------------------------------------
mv009	Forward Biobricking of r.BglII!
ccataGAATTCatgAGATCTatgaagattgatataacggactataaccatgc
mv011	Reverse Biobricking of r.BglII>
cgttaGGATCCatatgtcacgattgttcctc

Construction of {<r.BglII>} Basic Part K112108 

PCR mv012/mv011 on pBca9145-Bca1147    (697bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112108
-----------------------------------------------
mv012	Forward Biobricking of <r.BglII>
ccataGAATTCatgAGATCTaagattgatataacggactataacc
mv011	Reverse Biobricking of r.BglII>
cgttaGGATCCatatgtcacgattgttcctc

Construction of {rbs.ligase!} Basic Part K112109 

PCR mv013/mv016 on T4 DNA ligase    (269 bp, gp = A)
PCR mv015/mv018 on T4 DNA ligase    (376 bp, gp = B)
PCR mv017/mv020 on T4 DNA ligase    (191 bp, gp = C)
PCR mv019/mv014 on T4 DNA ligase    (746 bp, gp = D)
---------------------------------------------------
PCR mv013/mv018 on A+B              (622 bp, gp = E)
PCR mv017/mv014 on C+D              (914 bp, gp = F)
---------------------------------------------------
PCR mv013/mv014 on E+F              (1514bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                   (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112109
-----------------------------------------------
mv013	Forward Biobricking of rbs.ligase!
ccataGAATTCatgAGATCTaaactctaaaggatgaacaatg
mv014	Reverse Biobricking of ligase!
cgttaGGATCCtcatagaccagttacctcatg
mv015  Removing the 1st EcoRI site in T4 DNA ligase(F)  gacttcattgaGttcacattagc
mv016  Removing the 1st EcoRI site in T4 DNA ligase(R)  gctaatgtgaaCtcaatgaagtc
mv017  Removing the BglII site in T4 DNA ligase(F)  ctaggattagaCcttcttaagg
mv018  Removing the BglII site in T4 DNA ligase(R)  ccttaagaagGtctaatcctag
mv019  Removing the 2nd EcoRI site in T4 DNA ligase(F)  gctaaagaGttcgccgaagtagc
mv020  Removing the 2nd EcoRI site in T4 DNA ligase(R)  gctacttcggcgaaCtctttagc


Construction of {ligase!} Basic Part K112110 

PCR mv021/mv014 on pBca1256-K112109    (1495bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112110
-----------------------------------------------
mv021	Forward Biobricking of ligase!
ccataGAATTCatgAGATCTatgattcttaaaattctgaacg
mv014	Reverse Biobricking of ligase!
cgttaGGATCCtcatagaccagttacctcatg

Construction of {<ligase!} Basic Part K112111 

PCR mv022/mv014 on pBca1256-K112109    (1492bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112111
-----------------------------------------------
mv022	Forward Biobricking of <ligase!
ccataGAATTCatgAGATCTattcttaaaattctgaacg
mv014	Reverse Biobricking of ligase!
cgttaGGATCCtcatagaccagttacctcatg

Construction of {ligase>} Basic Part K112112 

PCR mv021/mv023 on pBca1256-K112109    (1492bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112112
-----------------------------------------------
mv021	Forward Biobricking of ligase!
ccataGAATTCatgAGATCTatgattcttaaaattctgaacg
mv023	Reverse Biobricking of ligase>
cgttaGGATCCtagaccagttacctcatg

Construction of {<ligase>} Basic Part K112113 

PCR mv022/mv023 on pBca1256-K112109    (1489bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112113
-----------------------------------------------
mv022	Forward Biobricking of <ligase!
ccataGAATTCatgAGATCTattcttaaaattctgaacg
mv023	Reverse Biobricking of ligase>
cgttaGGATCCtagaccagttacctcatg

Construction of {a~ligase!} Basic Part K112114 

PCR mv024/mv014 on pBca1256-K112109    (1499bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112114
-----------------------------------------------
mv024	Forward Biobricking of a~ligase!
ccataGAATTCatgAGATCTaacaatgattcttaaaattctgaacg
mv014	Reverse Biobricking of ligase!
cgttaGGATCCtcatagaccagttacctcatg

Construction of {a~Cre!} Basic Part K112115 

PCR mv025/mv002 on pBca9145-Bca1010    (1067bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112115
-----------------------------------------------
mv025	Forward Biobricking of a~Cre!
ccataGAATTCatgAGATCTttaaatgtccaatttactgaccgtacacc
mv002	Reverse Biobricking of <Cre!
cgttaGGATCCttaatcgccatcttccagc

Construction of {a~r.BamHI!} Basic Part K112116 

PCR mv026/mv006 on pBca9145-I716210    (677bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112116
-----------------------------------------------
mv026	Forward Biobricking of a~r.BamH1!
ccataGAATTCatgAGATCTtagaatggaagtagaaaaagagtttattactgatgaagc
mv006	Reverse Biobricking of r.BamH1!
cgttaGGATCCctatttgttttcaactttatctttcc

Construction of {a~r.BglII!} Basic Part K112117 

PCR mv027/mv010 on pBca9145-Bca1147    (707bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112117
-----------------------------------------------
mv027	Forward Biobricking of a~r.BglII!
ccataGAATTCatgAGATCTcataatgaagattgatataacggactataaccatgc
mv010	Reverse Biobricking of r.BglII!
cgttaGGATCCttaatatgtcacgattgttcctc

Construction of rrnB P1 promoter Basic Part K112118 

PCR mv028/mv029 on rrnB native sequence   (721bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                         (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112118
-----------------------------------------------
mv028	Forward Biobricking of rrnB P1 promoter
cgataGAATTCatgAGATCTatgcactaaaacgtatcctacgcccg
mv029   Reverse Biobricking of rrnB P1 promoter
cgttaGGATCCcgccttcccgctacagagtcaagc

Construction of {rbs.Cre!} Basic Part K112119 

PCR mv030/mv002 on pBca9145-Bca1010    (1082bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112119
-----------------------------------------------
mv030	Forward Biobricking of rbs.Cre!
cgataGAATTCatgAGATCTaccattatctgagtgttaaatgtccaatttactgaccgtacacc
mv002	Reverse Biobricking of <Cre!
cgttaGGATCCttaatcgccatcttccagc


Construction of {rbs.r.BamHI!} Basic Part K112120 

PCR mv031/mv006 on pBca9145-I716210    (692bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112120
-----------------------------------------------
mv031	Forward Biobricking of rbs.r.BamH1!
cgataGAATTCatgAGATCTtaagtgggggcaaattagaatggaagtagaaaaagagtttattactgatgaagc
mv006	Reverse Biobricking of r.BamH1!
cgttaGGATCCctatttgttttcaactttatctttcc

Construction of {rbs.r.BglII!} Basic Part K112121 

PCR mv032/mv010 on pBca9145-Bca1147    (722bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112121
-----------------------------------------------
mv032	Forward Biobricking of rbs.r.BglII!
cgataGAATTCatgAGATCTtggaaggagacactcataatgaagattgatataacggactataaccatgc
mv010	Reverse Biobricking of r.BglII!
cgttaGGATCCttaatatgtcacgattgttcctc

Construction of {Cre!} Basic Part K112122 

PCR mv003/mv002 on pBca9145-Bca1010    (1063bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112122
-----------------------------------------------
mv003	Forward Biobricking of Cre>
ccataGAATTCatgAGATCTatgtccaatttactgaccgtac
mv002	Reverse Biobricking of <Cre!
cgttaGGATCCttaatcgccatcttccagc

Construction of {<r.BglII!} Basic Part K112123 

PCR mv012/mv010 on pBca9145-Bca1147    (700bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                      (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112123
-----------------------------------------------
mv012	Forward Biobricking of <r.BglII>
ccataGAATTCatgAGATCTaagattgatataacggactataacc
mv010	Reverse Biobricking of r.BglII!
cgttaGGATCCttaatatgtcacgattgttcctc

Construction of {rbs.trfA!} Basic Part K112124 

PCR JH102F/JH102R on RP4 genome      (416bp, EcoRI/BamHI/DpnI)
Sub into pBjh1601KC                  (EcoRI/BamHI, 3132+778, L)
Product is pBjh1601KC-K112124
-----------------------------------------------
JH102F	 Forward Biobricking of {rbs.trfA!}
ccatgGAATTCatgAGATCTgcgaggggcggcttccgctAtgtacaaccag
JH102R	 Reverse Biobricking of {rbs.trfA!}
cgttaGGATCCtcagagccttccacgcagc

Construction of {OriV} Basic Part K112125 

PCR mv037/mv038 on pSMART Fosmid Blunt Vector (Lucigen)    (677bp, EcoRI/BamHI/DpnI)
Sub into pBca1256                        (EcoRI/BamHI, 2472+910, L)
Product is pBca1256-K112125
-----------------------------------------------
mv037  Forward biobricking of OriV from pSMART
cgataGAATTCatgAGATCTtgcttcctcgctcactcgac
mv038  Reverse biobricking of OriV form pSMART
gctagGGATCCaagccctcggctggttgc

Construction of {temp. sensitive promoter} Basic Part K112126 

PCR JH115F/JH115R on pAH57
sub into pBca1256
Product is pBca1256-K112126
-----------------------------------------------
JH115F  Forward biobricking of cI, Pr
cgataGAATTCatgAGATCTgaagattcttgctcaattgttatcagc
JH115R  Reverse biobricking of cI, Pr
gctagGGATCCagaattgatacaacctccttagtacatgc
Madhvi's Notebook
UC Berkeley Team Homepage
Retrieved from "http://2008.igem.org/Template:Team:UC_Berkeley/Notebook/MJV_construction"