From: Ethylene production with engineered Synechocystis sp PCC 6803 strains
Cloning parts, plasmids, strains and primers | Description | Source/remark |
---|---|---|
RBS a | Â | Â |
Rbs-O | AGGAGGACTAGCATG | [2] |
Rbs-34 | ATTAAAGAGGAGAAAACTAGCATG | [15] |
Rbs-30 | AAAGAGGAGAAAACTAGCATG | [15] |
Rbs-H | TAGTGGAGGTACTAGCATG | RBS* from [15] |
Rbs-C9 | AAAGGAGGTGATAGCATG | This study |
Rbs-C10 | AAAGGAGGTGATTAGCATG | This study |
Rbs-C11 | AAAGGAGGTGATCTAGCATG | This study |
Promoter | Â | Â |
Ptrc | TTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGAGCGGATAACAATTTCACAC | [2] |
3 × Ptrc | TTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGTTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGTTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGAGCGGATAACAATTTCACACA | This study |
5 × Ptrc | TTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGTTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGTTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGTTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGTTGACAATTAATCATCCGGCTCGTATAATGTGTGGAATTGTGAGCGGATAACAATTTCACACA | This study |
Terminator | Â | Â |
tt | Same as B0014 from http://www.partsregistry.org | [2] |
Plasmids | Â | Â |
pHKH001 | Integration vector disrupting slr0168 in Synechocystis genome with kanamycin resistance cassette a selection marker | [2] |
pHKH_BB | Bio-brick compatible derivative of pHKH001 carrying the Ptrc promoter and tt-transcription terminator | This study |
pHKH_rbsO_efe | Integration vector carrying efe under Ptrc promoter and Rbs-O | This study |
pHKH_rbs34_efe | Integration vector carrying efe under Ptrc promoter and Rbs-34 | This study |
pHKH_rbs30_efe | Integration vector carrying efe under Ptrc promoter and Rbs-30 | This study |
pHKH_rbsH_efe | Integration vector carrying efe under Ptrc promoter and Rbs-H | This study |
pHKH_rbsc9_efe | Integration vector carrying efe under Ptrc promoter and Rbs-C9 | This study |
pHKH_rbsc10_efe | Integration vector carrying efe under Ptrc promoter and Rbs-C10 | This study |
pHKH_rbsc11_efe | Integration vector carrying efe under Ptrc promoter and Rbs-C11 | This study |
pHKH_3xPtrc_efe | Integration vector carrying efe under 3xPtrc promoter and Rbs-30 | This study |
pHKH_5xPtrc_efe | Integration vector carrying efe under 5x Ptrc promoter and Rbs-30 | This study |
pVZ_LS | A bio-brick compatible version of mobilizable broad host self-replication plasmid pVZ321 containing the RSF1010 replicon, and carrying the Ptrc promoter, Rbs30 and tt-transcription terminator | Plasmid pVZ321 from [42] |
pVZ-efe | pVZ plasmid carrying efe gene under Ptrc promoter and rbs 30 | This study |
pVZ_Nhis-efe | pVZ plasmid carrying N-terminal hexa-histidine tagged efe under promoter Ptrc and Rbs-30 | This study |
Strains | Â | Â |
Wild-type (WT) | Synechocystis sp. PCC6803, glucose tolerant, naturally transformable | Bhaya (Stanford) |
VPV1 | WT, Slr0168::Ptrc-rbsO-efe-Kmr | This study |
VPV2 | WT, Slr0168::Ptrc-rbs34-efe-Kmr | This study |
VPV3 | WT, Slr0168::Ptrc-rbs30-efe-Kmr | This study |
VPV4 | WT, Slr0168::Ptrc-rbsH-efe-Kmr | This study |
VPV5 | WT, Slr0168::Ptrc-rbsc9-efe-Kmr | This study |
VPV6 | WT, Slr0168::Ptrc-rbsc10-efe-Kmr | This study |
VPV7 | WT, Slr0168::Ptrc-rbsc11-efe-Kmr | This study |
VPV40 | WT, Slr0168::3xPtrc-rbsO-efe-Kmr | This study |
VPV43 | WT, Slr0168::5xPtrc-rbsO-efe-Kmr | This study |
SAW11 | Glycogen knock out strain | [36] |
VPV55 | WT carrying plasmid pVZ _efe | This study |
VPV56 | SAW11 carrying plasmid pVZ _efe | This study |
VPV58 | SAW11 carrying plasmid pVZ_Nhis_efe | This study |
VPV62 | WT carrying plasmid pVZ_Nhis_efe | This study |
VPV65 | SAW11, Slr0168::Ptrc-rbs30-efe-Kmr | This study |
Primers | Â | Â |
efe-bb-fwd | GCGGAATTCGCGGCCGCTTCTAGAGGAGGACTAGCATGACCAACTTGCAAACCTTTGAAT | Â |
efe-bb-r | GTACTGCAGCGGCCGCTACTAGTATTAGGAGCCGGTGGCGCG | Â |
vpv-1-st-Hind-R | CTAGTAAAGCTTATCAATACTTTCCACCCC | Â |
VPV-efe-Bamh1-f-noatg | CCATCACGGATCCACCAACTTGCAAACCTTTGAATTGC | Â |
VPV-efe-st-Hind3-rev | TAATTAAGCTTATCAGGAGCCGGTGGCGCGGGTATCGG | Â |
V-rbs30-efe-f | GACAGCTAGCATTAAAGAGGAGAAAACTAGCATGACCAACTTGCAAACC | Â |
V-rbs34-efe-f | GACAGCTAGCAAAGAGGAGAAAACTAGCATGACCAACTTGCAAACC | Â |
V-rbsC10-efe-f | GACAGCTAGCAAAGGAGGTGATTAGCATGACCAACTTGCAAACC | Â |
V-rbsC11-efe-f | GACAGCTAGCAAAGGAGGTGATCTAGCATGACCAACTTGCAAACC | Â |
V-rbsC9-efe-f | GACAGCTAGCAAAGGAGGTGATAGCATGACCAACTTGCAAACC | Â |
V-rbsH-efe-f | GACAGCTAGCTAGTGGAGGTACTAGCATGACCAACTTGCAAACC | Â |
v-bbs-pUC-r | CGGGATCCGATCCAATCTGCAGCGGCCGCTACTAGTA | Â |
H1-SEG-VF | TGTCGCCGCTAAGTTAGACCGC | Â |
H2-SEG-VR | CTGTGGGTAGTAAACTGGCAATGCC | Â |
H1-SEQ-VF | CGGCAATGGTCCCAAAAT | Â |
KanF-SEQ-VR | AGACGTTTCCCGTTGAAT | Â |
pVZ321-SEQ-F | CGCAGGGCTTTATTGATT | Â |
pVZ321-SEQ-R | CCCCCCCCACTCTATTGTA | Â |
efe-n–H-f | CAGATATGACATATGCATCATCATCATCATCATACCAACTTGCAAACCTTTGAATTGC |  |
efe-ba-r | ACCTAGGTCAGTAGGATCCTTATTAGGAGCCGGTGGCGCGGGTATCG | Â |