Paris/July 21

[[Paris/July 20|yesterday]] -- [[Paris/July 22|tomorrow]] <br>
== Digestion reactions ==

Double digestion reactions were performed as follows:
for a double digestion by X & Y, the template was digested by X(mix1:in a 25µL reaction mix)& Y (mix2: in a 25µL reaction mix) for 2h at 37°. Then, enzyme Y was added to mix1 & enz. X added to mix 2. This second reaction was also performed at 37° for 2h.


25µL reaction mix:
*10µL DNA template (miniprep or PCR product)
*1µL enz. (added last)
*2.5µL 10x reaction buffer (depending on enzymes used)
*BSA 100x 0.25µL
*H2O qsp 25µL


{|
|- style="background: #ccccff;"
! colspan="5" style="background: #ccccff;" | Digestion Products
|- style="background: #ccccff; text-align: center;"
|width=5%| Number
|width=20%| Product Name
|width=30%| Matrix Name
|width=5%| Enzyme 1
|width=5%| Enzyme 2
|width=5%| size
|width=35%| Description
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D9
|<bbpart>pJ23100</bbpart> ES vector
|<bbpart>pJ23100</bbpart> (MP1.1 & MP1.2)
|EcoI
|SpeI
|2096
|Vector with mRFP RBS/CDS ready for insertion promoter (fw-insert)
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D10
|<bbpart>pJ23100</bbpart> promoter fw-insert
|<bbpart>pJ23100</bbpart> (MP1.1 & MP1.2)
|EcoI
|SpeI
|58
|J23100 promoter ready for use as a fw-insert
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D11
|<bbpart>pJ23107</bbpart> ES vector
|<bbpart>pJ23107</bbpart> (MP2.1, MP2.2, MP2.3 & MP2.4)
|EcoRI
|SpeI
|2096
|Vector with mRFP RBS/CDS ready for insertion of a fw-insert promoter
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D12
|<bbpart>pJ23107</bbpart> promoter fw-insert
|<bbpart>pJ23107</bbpart> (MP2.1, MP2.2, MP2.3 & MP2.4)
|EcoRI
|SpeI
|58
|J23107 promoter ready for usage as a fw-insert
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D13
|<bbpart>mRFP</bbpart> 
|<bbpart>pJ23107</bbpart> (MP2.2, MP2.3 & MP2.4)
|SpeI
|PstI
|887
|mRFP RBS/CDS/T sequence, can it be used as a backward insert? => yes but you will not be able to add any biobrick after it
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D14
|Cre ORF 
|<bbpart>J61047</bbpart> MP9.1 & MP9.2
|XbaI
|PstI
|1063
|Cre ORF extracted for usage as a bw-insert
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D15
|<bbpart>B0030</bbpart> bw-vector
|<bbpart>B0030</bbpart> MP5.1 & MP5.2
|SpeI
|PstI
|2076
|bacward vector for cloning a CDS bw-insert after the B0030 RBS
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D16
|<bbpart>pSB1AK3</bbpart> vector
|<bbpart>B0015</bbpart> MP3.1 & MP3.2
|EcoRI
|PstI
|3148
|pSB1AK3 open vector for BioBrick cloning
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D17
|pSB1A2 <bbpart>E0422</bbpart> vector
|<bbpart>E0422</bbpart> (MP6.1 & MP6.2)
|EcoRI
|PstI
|2038
|pSB1A2 open vector for BioBrick cloning
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D18
|BB dig. lox66DapAE.coli
|lox66-DapAE.coli P5 PCR product
|EcoRI
|PstI
|~960
|PCR product digested for cloning as a BioBrick
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D19
|BB dig. lox66DapAsubtilis
|lox66-DapAsubtilis P6 PCR product
|EcoRI
|PstI
|~960
|PCR product digested for cloning as a BioBrick
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D20
|fw insert lox66-DapAE.coli
|lox66-DapAE.coli P5 PCR product
|XbaI
|PstI
|~960
|PCR product digested for usage as a forward insert
|
|
|- style="background: #cccccc;"  
| style="background: #ccffcc;" |D21
|fw insert lox66-DapAsubtilis
|lox66-DapAsubtilis P6 PCR product
|XbaI
|PstI
|~960
|PCR product digested for usage as a forward insert
|
|
|}