LOCUS       sequence-387246-        6057 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression of Thiolredoxin-3C cleavage site-CarHCN.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6057)
  AUTHORS   Yang Z, Fok HKF, Luo J, Yang Y, Wang R, Huang X, Sun F
  TITLE     B(12)-induced reassembly of split photoreceptor protein enables 
            photoresponsive hydrogels with tunable mechanics.
  JOURNAL   Sci Adv. 2022 Apr;8(13):eabm5482. doi: 10.1126/sciadv.abm5482. Epub 
            2022 Apr 1.
  PUBMED    35363531
REFERENCE   2  (bases 1 to 6057)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2022-04"; volume: "8(13):eabm5482. doi"; pages: " 
            10.1126/sciadv.abm5482. Epub 2022 Apr 1"
COMMENT     Sequence Label: p32m3C Trx-CN
FEATURES             Location/Qualifiers
     source          1..6057
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      200..788
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     689..708
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     942..959
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    974..1113
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1039..1058)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1199..1221
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1215..1406)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2215..3297)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     3259..3278
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3298..3375)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3581..3600
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3688..3707
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3688..3706
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3707..3731
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             3746..3768
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3776..4102
                     /codon_start=1
                     /gene="trxA"
                     /product="E. coli thioredoxin"
                     /label=TrxA
                     /translation="MSDKIIHLTDDSFDTDVLKADGAILVDFWAEWCGPCKMIAPILDE
                     IADEYQGKLTVAKLNIDQNPGTAPKYGIRGIPTLLLFKNGEVAATKVGALSKGQLKEFL
                     DANLA"
     CDS             4124..4141
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4151..4174
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4472..4489
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4542..4560)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4556..4603
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4640..5095
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4727..4746)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4937..4958
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5121..5225
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5226..5294,5295..6057,1..29)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5444..5463)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
       61 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
      121 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
      181 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
      241 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
      301 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
      361 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
      421 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
      481 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
      541 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
      601 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
      661 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
      721 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
      781 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
      841 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
      901 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
      961 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     1021 gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact
     1081 ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac
     1141 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     1201 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg
     1261 taaagctcat cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc
     1321 agctcgttga gtttctccag aagcgttaat gtctggcttc tgataaagcg ggccatgtta
     1381 agggcggttt tttcctgttt ggtcactgat gcctccgtgt aagggggatt tctgttcatg
     1441 ggggtaatga taccgatgaa acgagagagg atgctcacga tacgggttac tgatgatgaa
     1501 catgcccggt tactggaacg ttgtgagggt aaacaactgg cggtatggat gcggcgggac
     1561 cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta atacagatgt aggtgttcca
     1621 cagggtagcc agcagcatcc tgcgatgcag atccggaaca taatggtgca gggcgctgac
     1681 ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag
     1741 gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc
     1801 tgctaaccag taaggcaacc ccgccagcct agccgggtcc tcaacgacag gagcacgatc
     1861 atgcgcaccc gtggggccgc catgccggcg ataatggcct gcttctcgcc gaaacgtttg
     1921 gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca agattccgaa taccgcaagc
     1981 gacaggccga tcatcgtcgc gctccagcga aagcggtcct cgccgaaaat gacccagagc
     2041 gctgccggca cctgtcctac gagttgcatg ataaagaaga cagtcataag tgcggcgacg
     2101 atagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc
     2161 ggtcgagatc ccggtgccta atgagtgagc taacttacat taattgcgtt gcgctcactg
     2221 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     2281 gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg
     2341 caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct
     2401 ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga
     2461 gctgtcttcg gtatcgtcgt atcccactac cgagatgtcc gcaccaacgc gcagcccgga
     2521 ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt
     2581 gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca
     2641 gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc
     2701 agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg
     2761 gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat
     2821 aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca
     2881 ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact
     2941 gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc
     3001 taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac
     3061 aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg
     3121 tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc
     3181 ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac
     3241 ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt
     3301 caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg
     3361 ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc
     3421 agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg
     3481 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag
     3541 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg
     3601 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga
     3661 tcgagatcga tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg
     3721 ataacaattc ccctctagaa ataattttgt ttaactttaa gaaggagata tacatatgag
     3781 cgataaaatt attcacctga ctgacgacag ttttgacacg gatgtactca aagcggacgg
     3841 ggcgatcctc gtcgatttct gggcagagtg gtgcggtccg tgcaaaatga tcgccccgat
     3901 tctggatgaa atcgctgacg aatatcaggg caaactgacc gttgcaaaac tgaacatcga
     3961 tcaaaaccct ggcactgcgc cgaaatatgg catccgtggt atcccgactc tgctgctgtt
     4021 caaaaacggt gaagtggcgg caaccaaagt gggtgcactg tctaaaggtc agttgaaaga
     4081 gttcctcgac gctaacctgg ccggttctgg ttctggccat atgcaccatc atcatcatca
     4141 ttcttctggt ctggaagttc tgttccaggg gcccggatcc gaattcccag aagatctggg
     4201 caccggcctg ctggaagcac tgctgcgcgg tgatctggcg ggcgccgaag ctctgtttcg
     4261 tcgtggcctg cgtttctggg gcccggaagg cgttctggag cacctgctgc tgccggtgct
     4321 gcgtgaagtg ggcgaggctt ggcaccgtgg tgaaatcggc gttgcagaag aacacctggc
     4381 gagcaccttc ctgcgcgcgc gtctgcagga gctgctggac ctggcaggtt tcccgccggg
     4441 tccgccgtaa aagcttgcgg ccgcactcga gcaccaccac caccaccact gagatccggc
     4501 tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc
     4561 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat
     4621 atccggattg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4681 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4741 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4801 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     4861 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     4921 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     4981 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     5041 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
     5101 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5161 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5221 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5281 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5341 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5401 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5461 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5521 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5581 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5641 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5701 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5761 cgatgcctgc agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5821 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5881 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     5941 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6001 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgaga
//
