BBa_K2009430

BBa_K2009430 Version 1

Component

Source:
http://parts.igem.org/Part:BBa_K2009430
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Yin Wu
Date created: 2016-09-13 11:00:00
Date modified: 2016-09-17 07:33:46

sfGFP1-10



Types
DnaRegion

Roles
CDS

Coding

Sequences BBa_K2009430_sequence (Version 1)

Description

sfGFP1-10??????PSB1A3 length: 642bp
Derived from: PCR from Part:BBa_I746916,Cambridge 2008

sfGFP1-10??????PSB1A3 is an expression plasmid which insertsfGFP1-10 into PSB1A3.

PSB1A3 is a high copy numberplasmid carrying ampicillin resistance. The replication origin is aPUC19-derived pMB1.

SfGFP1-10 is a part of GFP(from 1bp to 214bp), GFP has been mutated to improve its solubilityand self-associating activity. When it express, it will emit green fluorescenceslightly under the fluorescence microscope.

We try to find anideal protein tag to be work both invivo and invitro and it can provide a sensitive measurable signalwhich don???t need external chemical reagents or substrates. Finally we find away to accomplish this goal?????? dividing GFP into sfGFP1-10and sfGFP11. Either the sfGFP1-10 or sfGFP11 will emit green fluorescence slightly under the fluorescencemicroscope. However, when sfGFP1-10 and sfGFP11 express insame cell, they will interact each other and emit more intense fluorescence thaneach of them. The split GFP system is simple and does not change fusion proteinsolubility.

Primers for these biobrickvectors can be found in part: BBa_G00100 (aka VF2) and part: BBa_G00101 (akaVR)

Usage and biology:The split GFP system has manypractical applications. Obtaining soluble, well-folded recombinant proteins fordownstream applications requires screening large numbers of protein variants (mutants,fragments, fusion tags, folding partners) and testing many expression orrefolding conditions.(Ste??phanieCabantous, Thomas C Terwilliger & Geoffrey S Waldo,2005)

Part sequence

atgcgtaaaggcgaagagctgttcactggtgtcgtccctattctggtggaactggatggtgatgtcaacggtcataagttttccgtgcgtggcgagggtgaaggtgacgcaactaatggtaaactgacgctgaagttcatctgtactactggtaaactgccggtaccttggccgactctggtaacgacgctgacttatggtgttcagtgctttgctcgttatccggaccatatgaagcagcatgacttcttcaagtccgccatgccggaaggctatgtgcaggaacgcacgatttcctttaaggatgacggcacgtacaaaacgcgtgcggaagtgaaatttgaaggcgataccctggtaaaccgcattgagctgaaaggcattgactttaaagaagacggcaatatcctgggccataagctggaatacaattttaacagccacaatgtttacatcaccgccgataaacaaaaaaatggcattaaagcgaattttaaaattcgccacaacgtggaggatggcagcgtgcagctggctgatcactaccagcaaaacactccaatcggtgatggtcctgttctgctgccagacaatcactatctgagcacgcaaagcgttctgtctaaagatccgaacgagaaa
(All the sequence has been testified by Sangon)

assume protein structure

We used Phyre2 to get the assume structure:

图片名称

(by PyMOL)

Citation:

The Phyre2 web portal for protein modeling, prediction and analysis

Kelley LA et al.

Nature Protocols 10, 845-858 (2015).

Notes

We PCR from Part:BBa_I746916,Cambridge 2008 and standardize it.

Source

PCR from Part:BBa_I746916,Cambridge 2008.

Sequence Annotation Location Component / Role(s)
sfGFP1-10
1,642
CDS feature/protein
igem#experience
None
 
igem#status
Planning
 
synbiohub#ownedBy
user/james
 
synbiohub#ownedBy
user/myers
 
synbiohub#topLevel
BBa_K2009430/1