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Team:Harvard/flavor/notebook
From 2010.igem.org
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| + | ===Primer Designs for Agrobacterium Vector=== | ||
| + | *Primers were designed to amplify from the Expression Series pORE Agrobacterium plasmid (e3) 1)the pENTcup2 promoter, 2) the tNOS stop sequence, 3) the tNOS stop sequence + additional stop codon and 4) the pHLP promoter.<br> | ||
| + | Note: the NOSterm_BB_R primer works for both tNOS and tNOS+STOP codon sequences. | ||
| + | pENTcup2_BB_F | ||
| + | CCTTTCTAGAGGGATCTTCTGCAAGCATCT | ||
| + | pENTcup2_BB_R | ||
| + | AAGGCTGCAGCGGCCGCTACTAGTTCCGGTGGGTTTTGAGGT | ||
| + | |||
| + | STOP_NOSterm_BB_F | ||
| + | CCTTTCTAGATGAGATCGTTCAAACATTTGG | ||
| + | |||
| + | NOSterm_BB_F | ||
| + | CCTTTCTAGAGATCGTTCAAACATTTGGCA | ||
| + | |||
| + | NOSterm_BB_R | ||
| + | AAGGCTGCAGCGGCCGCTACTAGTGATCTAGTAACATAGATGACA | ||
| + | |||
| + | pHPL_BB_F | ||
| + | CCTTTCTAGAAACGTGGATACTTGGCAGTG | ||
| + | |||
| + | pHPL_BB_R | ||
| + | AAGGCTGCAGCGGCCGCTACTAGTCTTTTGAGCTTAGAGGTTTTT | ||
| + | |||
| + | ===BioBrick Transformation=== | ||
| + | *Colonies were observed after overnight culture growth, but in low quantities. | ||
| + | **J45700 and J45004 (both Wintergreen Pathway) showed minimal number of colonies on both 10μL and 100μL cultures. | ||
| + | **J45250 and J45014 (both Banana Pathway) showed no colonies on either the 10μL or 100μL cultures. | ||
| + | |||
| + | ===Codon Usage in Arabidopsis=== | ||
| + | Using http://gcua.schoedl.de/ <br><br> | ||
| + | '''Wintergreen Pathway:''' <br> | ||
| + | BioBrick Part #: BBa_J45700 <br> | ||
| + | Original Organism: ''Pseudomonas aeruginosa'' <br> | ||
| + | |||
| + | [[Image:Wintergreen Pathway (full) Compare_Page_1.jpg|150px|BBa_J45700]] | ||
| + | [[Image:Wintergreen Pathway (full) Compare_Page_2.jpg|150px|BBa_J45700]] | ||
| + | [[Image:Wintergreen Pathway (full) Compare_Page_3.jpg|150px|BBa_J45700]] | ||
| + | [[Image:Wintergreen Pathway (full) Compare_Page_4.jpg|150px|BBa_J45700]] | ||
| + | |||
| + | '''Banana Pathway:''' <br> | ||
| + | BioBrick Part #: BBa_J45014 <br> | ||
| + | Original Organism: ''Saccharomyces cerevisiae'' <br> | ||
| + | [[Image:codon_table_BBa_J45014.jpg|150px|BBa_J45014]] | ||
| + | [[Image:codon_table_BBa_J45014_pg_2.jpg|150px|BBa_J45014]] | ||
| + | [[Image:Codon_table_BBa_J45014_pg_3.jpg|150px|BBa_J45014]] | ||
| + | |||
| + | BioBrick Part #: BBa_J45250 <br> | ||
| + | [[Image:codon_table_BBa_J45250_pg1.jpg|150px|BBa_J45250]] | ||
| + | [[Image:codon_table_BBa_J45250_pg2.jpg|150px|BBa_J45250]] | ||
| + | [[Image:codon_table_BBa_J45250_pg3.jpg|150px|BBa_J45250]] | ||
| + | [[Image:codon_table_BBa_J45250_pg4.jpg|150px|BBa_J45250]] | ||
| + | |||
| + | '''Valencene Pathway:''' | ||
| + | |||
| + | [[Image:Citrus_sinesis_valencene_synthase_Page_1.jpg|150px|Valencene]] | ||
| + | [[Image:Citrus_sinesis_valencene_synthase_Page_2.jpg|150px|Valencene]] | ||
| + | [[Image:Citrus_sinesis_valencene_synthase_Page_3.jpg|150px|Valencene]] | ||
| + | |||
| + | ===Valencene Extraction=== | ||
| + | Used RNeasy Plant Mini Kit to extract RNA from the <u>flavedo</u> of an Organic Valencia Orange. | ||
| + | |||
| + | [[Image:Flavedo.png|500px|Flavedo]] | ||
==<html><a class="labnotebook" name="06-16-2010">06-16-2010</a></html> [ [[#top|top]] ]== | ==<html><a class="labnotebook" name="06-16-2010">06-16-2010</a></html> [ [[#top|top]] ]== | ||
Revision as of 00:19, 25 October 2010
notebook calendar [ top ]
06-14-2010 [ top ]
- Miraculin and brazzein constructs due to arrive on Wednesday from Mr. Gene
BioBrick Transformation
BioBrick parts from the 2010 iGEM kit were transformed and grown in highly competent TURBO bacteria.
- Wintergreen Scent Pathway:
BBa_J45700 - entire pathway, Ampicillin
BBa_J45004 - BSMT1 only, Ampicillin
(Not in 2010 BB Kit: BBa_J45017 - PchB, PchA)
- Banana Scent Pathway:
BBa_J45250 - ATF3 + Promoter, Ampicillin?
BBa_J45014 - ATF3 only, Ampicillin
(Not in BB 2010 Kit: BBa_J45400 - BAT2 and THI3)
06-15-2010 [ top ]
Primer Designs for Agrobacterium Vector
- Primers were designed to amplify from the Expression Series pORE Agrobacterium plasmid (e3) 1)the pENTcup2 promoter, 2) the tNOS stop sequence, 3) the tNOS stop sequence + additional stop codon and 4) the pHLP promoter.
Note: the NOSterm_BB_R primer works for both tNOS and tNOS+STOP codon sequences.
pENTcup2_BB_F CCTTTCTAGAGGGATCTTCTGCAAGCATCT
pENTcup2_BB_R AAGGCTGCAGCGGCCGCTACTAGTTCCGGTGGGTTTTGAGGT
STOP_NOSterm_BB_F CCTTTCTAGATGAGATCGTTCAAACATTTGG
NOSterm_BB_F CCTTTCTAGAGATCGTTCAAACATTTGGCA
NOSterm_BB_R AAGGCTGCAGCGGCCGCTACTAGTGATCTAGTAACATAGATGACA
pHPL_BB_F CCTTTCTAGAAACGTGGATACTTGGCAGTG
pHPL_BB_R AAGGCTGCAGCGGCCGCTACTAGTCTTTTGAGCTTAGAGGTTTTT
BioBrick Transformation
- Colonies were observed after overnight culture growth, but in low quantities.
- J45700 and J45004 (both Wintergreen Pathway) showed minimal number of colonies on both 10μL and 100μL cultures.
- J45250 and J45014 (both Banana Pathway) showed no colonies on either the 10μL or 100μL cultures.
Codon Usage in Arabidopsis
Using http://gcua.schoedl.de/
Wintergreen Pathway:
BioBrick Part #: BBa_J45700
Original Organism: Pseudomonas aeruginosa
BBa_J45700 BBa_J45700 BBa_J45700 BBa_J45700
Banana Pathway:
BioBrick Part #: BBa_J45014
Original Organism: Saccharomyces cerevisiae
BBa_J45014
BBa_J45014
BBa_J45014
BioBrick Part #: BBa_J45250
BBa_J45250
BBa_J45250
BBa_J45250
BBa_J45250
Valencene Pathway:
Valencene Extraction
Used RNeasy Plant Mini Kit to extract RNA from the flavedo of an Organic Valencia Orange.
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_Compare_Page_4.jpg){kind=link}
