Difference between revisions of "Part:BBa K596004"

Latest revision as of 16:17, 10 May 2013

Algae Protein Expression Vector

This Biobrick Plasmid was created from pRbcRL (which is also derived from pBluescriptKS-) by putting biobrick prefix and suffix between HSP70A/RbcS2 promoter and 3'UTR sequences from RbcS2 gene. We used pRbcRL(Hsp196)for the backbone plasmid. We removed luciferase protein coding sequence(which is abbreviated as crluc)using XhoI/BamHI restriction. Then we ligated with excised plasmid with standard Biobrick prefix and suffix. Biobrick prefix and suffix contains four restriction enzymes namely XbaI, EcoRI, SpeI, PstI. This plasmid can be used for protein expression in C. reinhardtii and for cloning purposes in E.coli. it contains ampicillin for selection in E.coli. F1 origin of replication is for double stranded replication process. HSP70A is a sequence for transcriptional activation that enhances expression of the gene.

References:

1. Schroda M, Blöcker D, Beck CF. The HSP70A promoter as a tool for the improved expression of transgenes in Chlamydomonas. The Plant journal : for cell and molecular biology. 2000;21(2):121-31. Available at: http://www.ncbi.nlm.nih.gov/pubmed/10743653.

2. Fuhrmann M, Hausherr A, Ferbitz L, et al. Monitoring dynamic expression of nuclear genes in Chlamydomonas reinhardtii by using a synthetic luciferase reporter gene. Plant molecular biology. 2004;55(6):869-81. Available at: http://www.ncbi.nlm.nih.gov/pubmed/15604722.

Map:

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal prefix found in sequence at 3787
Illegal suffix found in sequence at 1
12
INCOMPATIBLE WITH RFC[12]
Illegal EcoRI site found at 3787
Illegal NheI site found at 3376
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NotI site found at 9
Illegal NotI site found at 3793
21
INCOMPATIBLE WITH RFC[21]
Illegal EcoRI site found at 3787
Illegal BamHI site found at 23
Illegal XhoI site found at 3779
23
INCOMPATIBLE WITH RFC[23]
Illegal prefix found in sequence at 3787
Illegal suffix found in sequence at 2
25
INCOMPATIBLE WITH RFC[25]
Illegal prefix found in sequence at 3787
Illegal XbaI site found at 3802
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NgoMIV site found at 2598
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 1621
Illegal SapI site found at 538

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K596004 short</partinfo>
-This Biobrick Plasmid is created by putting biobrick prefix and suffix between HSP70/RBSC2 promoter and 3'UTR.
+This Biobrick Plasmid was created from pRbcRL (which is also derived from pBluescriptKS-) by putting biobrick prefix and suffix between HSP70A/RbcS2 promoter and 3'UTR sequences from RbcS2 gene.
-We used pRbcRL(Hsp196)for bacbone plasmid. We excised luciferase gene(crluc)using XhoI/BamHI restriction. Then we ligated with biobrick prefix and suffix. This plasmid can be used for protein expression in C.Reinhardtii.
+We used pRbcRL(Hsp196)for the backbone plasmid. We removed luciferase protein coding sequence(which is abbreviated as crluc)using XhoI/BamHI restriction. Then we ligated with excised plasmid with standard Biobrick prefix and suffix. Biobrick prefix and suffix contains four restriction enzymes namely XbaI, EcoRI, SpeI, PstI. This plasmid can be used for protein expression in <i>C. reinhardtii </i> and for cloning purposes in <i>E.coli</i>. it contains ampicillin for selection in E.coli. F1 origin of replication is for double stranded replication process.
-HSP70 is a transcriptional activator that enhances expression of a gene.
+HSP70A is a sequence for transcriptional activation that enhances expression of the gene.
 '''References:'''
 . Schroda M, Blöcker D, Beck CF. The HSP70A promoter as a tool for the improved expression of transgenes in Chlamydomonas. The Plant journal : for cell and molecular biology. 2000;21(2):121-31. Available at: http://www.ncbi.nlm.nih.gov/pubmed/10743653.
 . Fuhrmann M, Hausherr A, Ferbitz L, et al. Monitoring dynamic expression of nuclear genes in Chlamydomonas reinhardtii by using a synthetic luciferase reporter gene. Plant molecular biology. 2004;55(6):869-81. Available at: http://www.ncbi.nlm.nih.gov/pubmed/15604722.