Difference between revisions of "Part:BBa K864444"

Latest revision as of 16:28, 29 September 2012

RFP-Linker-SYFP2

This part is a component of the Uppsala University iGEM 2012 modular small RNA screening system, together with the spot42 sRNA BBa_K864440. It can also be used for studing up- or downregulation of genes by other mechanisms.

Use this system to screen for downregulation of your gene by a random small RNA library. PCR amplify your 5’UTR, adding XbaI and EcoRI sites upstream and a BamHI site downstream, and clone into any backbone carrying this part, replacing the RFP cassette. By cloning with EcoRI and BamHI, the cloning will function as a red/white screening system. You have now created a novel BioBrick, that can be used to screen for translational downregulation of your gene of interest.

SYFP2 (BBa_K864100) is situated downstream of the RFP cassette BBa_J04450 and is connected in frame with a 10 aa glycine-serine repeated linker (BBa_J18922), to prevent folding errors in the SYFP2. Before the linker there is a BamHI restriction site (GGATCC), which translates into glycine-serine in E coli.

When screening for working small RNAs, it's recommended to include the first 15 codons of your gene of interest.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 1070
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 781
Illegal AgeI site found at 893
1000
COMPATIBLE WITH RFC[1000]

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K864444 short</partinfo>
-Use this part to insert any 5’ÚTR of your gene of interest to screen for downregulation of your gene with small RNA. PCR amplify your 5’UTR with EcoRI and BamHI and clone into any backbone carrying this part. Digest whis part with EcoRI and BamHI and as RFP is replaced, this part functions as a red/white screening system. This allows you to create a reporter system with a fluorescent marker for screening for functional small RNAs.
+This part is a component of the Uppsala University iGEM 2012 modular small RNA screening system, together with the spot42 sRNA <partinfo>K864440</partinfo>. It can also be used for studing up- or downregulation of genes by other mechanisms.
-SYFP2 <partinfo>BBa_K864100</partinfo> is situated downstrean of the RFP <partinfo>BBa_J04450</partinfo> and is connected in frame with a 12 aa glycine-serine repeated linker <partinfo>BBa_J18922</partinfo>, to prevent folding errors in the YFP or the 5’UTR. The first 6 nt of the glycine-serine linker is a BamHI restriction site (GGATCC), which translates into glycine-serine in E.Coli.
+Use this system to screen for downregulation of your gene by a random small RNA library. PCR amplify your 5’UTR, adding XbaI and EcoRI sites upstream and a BamHI site downstream, and clone into any backbone carrying this part, replacing the RFP cassette. By cloning with EcoRI and BamHI, the cloning will function as a red/white screening system. You have now created a novel BioBrick, that can be used to screen for translational downregulation of your gene of interest.
-When screening for working small RNAs its recommended to include the first 15 codons of your gene of interest.
+SYFP2 (<partinfo>BBa_K864100</partinfo>) is situated downstream of the RFP cassette <partinfo>BBa_J04450</partinfo> and is connected in frame with a 10 aa glycine-serine repeated linker (<partinfo>BBa_J18922</partinfo>), to prevent folding errors in the SYFP2. Before the linker there is a BamHI restriction site (GGATCC), which translates into glycine-serine in E coli.
+When screening for working small RNAs, it's recommended to include the first 15 codons of your gene of interest.
 <!-- Add more about the biology of this part here