LOCUS       sequence-322530-        4802 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Improved PhoCl variant with faster dissociation rate.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4802)
  AUTHORS   Lu X, Wen Y, Zhang S, Zhang W, Chen Y, Shen Y, Lemieux MJ, Campbell 
            RE
  TITLE     Photocleavable proteins that undergo fast and efficient 
            dissociation.
  JOURNAL   Chem Sci. 2021 May 31;12(28):9658-9672. doi: 10.1039/d1sc01059j. 
            eCollection 2021 Jul 21.
  PUBMED    34349937
REFERENCE   2  (bases 1 to 4802)
  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"; doi: 
            "10.1039/d1sc01059j"; journalName: "Chem Sci"; date: "2021-05-31- 
            31"; volume: "12"; issue: "28"; pages: "9658-9672"
COMMENT     Sequence Label: pBad-PhoCl2f
FEATURES             Location/Qualifiers
     source          1..4802
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             331..348
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             352..384
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(388..396,397..411)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             430..1155
                     /codon_start=1
                     /product="photocleavable protein that separates into two 
                     fragments upon exposure to violet light (Zhang et al., 
                     2017)"
                     /label=PhoCl
                     /translation="VIPDYFKQSFPEGYSWERSMTYEDGGICIATNDITMEGDSFINKI
                     HFQGTNFPPNGPVMQKRTVGWEASTEKMYERDGVLKGDVKMKLLLKGGGHYRCDYRTTY
                     KVKQKPVKLPDCHFVDHRIEILSHDKDYNKVKLYEHAVAKTSTDSMDELYKGGSGGMVS
                     KGEETITSVIKPDMKNKLRMEGNVNGHAFVIEGEGSGKPFEGSQTIDLEVKEGAPLPFA
                     YDILTTAFHYGNRVFTKYPR"
     primer_bind     complement(1203..1220)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1373..1459
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1551..1578
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1597..1688
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1689..1757,1758..2549)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1907..1926)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2720..3308
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3209..3228
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3462..3479
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3494..3634
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3560..3579)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3720..3742
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3898..4776)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
      301 taacaggagg aattaaccat ggggggttct catcatcatc atcatcatgg tatggctagc
      361 atgactggtg gacagcaaat gggtcgggat ctgtacgacg atgacgataa ggatccgagc
      421 tcgaggatgg tgatccctga ctacttcaag cagagcttcc ccgagggcta cagctgggag
      481 cgcagcatga cctacgagga cggcggcatc tgcatcgcca ccaacgacat cacaatggag
      541 ggggacagct tcatcaacaa gatccacttc cagggcacga acttcccccc caacggcccc
      601 gtgatgcaga agaggaccgt gggctgggag gccagcaccg agaagatgta cgagcgcgac
      661 ggcgtgctga agggcgacgt gaagatgaag ctgctgctga agggcggcgg ccactatcgc
      721 tgcgactacc gcaccaccta caaggtcaag cagaagcccg taaagctgcc cgactgccac
      781 ttcgtggacc accgcatcga gatcctgagc cacgacaagg actacaacaa ggtgaagctg
      841 tacgagcacg ccgtggccaa gacttccacc gacagcatgg acgagctgta caagggtggc
      901 agcggtggca tggtgagcaa gggcgaggag accattacaa gcgtgatcaa gcctgacatg
      961 aagaacaagc tgcgcatgga gggcaacgtg aacggccacg ccttcgtgat cgagggcgag
     1021 ggcagcggca agcccttcga gggctctcag acgattgatt tggaggtgaa ggagggcgcc
     1081 ccgctgccct tcgcctacga catcctgacc accgccttcc actacggcaa ccgcgtgttc
     1141 accaagtacc cacggtaagg taccaagctt ggctgttttg gcggatgaga gaagattttc
     1201 agcctgatac agattaaatc agaacgcaga agcggtctga taaaacagaa tttgcctggc
     1261 ggcagtagcg cggtggtccc acctgacccc atgccgaact cagaagtgaa acgccgtagc
     1321 gccgatggta gtgtggggtc tccccatgcg agagtaggga actgccaggc atcaaataaa
     1381 acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc
     1441 tctcctgagt aggacaaatc cgccgggagc ggatttgaac gttgcgaagc aacggcccgg
     1501 agggtggcgg gcaggacgcc cgccataaac tgccaggcat caaattaagc agaaggccat
     1561 cctgacggat ggcctttttg cgtttctaca aactcttttg tttatttttc taaatacatt
     1621 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     1681 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     1741 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     1801 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     1861 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     1921 tattatcccg tgttgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     1981 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     2041 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     2101 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     2161 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     2221 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     2281 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     2341 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     2401 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     2461 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     2521 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     2581 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     2641 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     2701 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     2761 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     2821 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     2881 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     2941 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     3001 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     3061 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     3121 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     3181 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     3241 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     3301 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     3361 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     3421 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     3481 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     3541 gcatatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     3601 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     3661 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     3721 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagcagat
     3781 caattcgcgc gcgaaggcga agcggcatgc ataatgtgcc tgtcaaatgg acgaagcagg
     3841 gattctgcaa accctatgct actccgtcaa gccgtcaatt gtctgattcg ttaccaatta
     3901 tgacaacttg acggctacat cattcacttt ttcttcacaa ccggcacgga actcgctcgg
     3961 gctggccccg gtgcattttt taaatacccg cgagaaatag agttgatcgt caaaaccaac
     4021 attgcgaccg acggtggcga taggcatccg ggtggtgctc aaaagcagct tcgcctggct
     4081 gatacgttgg tcctcgcgcc agcttaagac gctaatccct aactgctggc ggaaaagatg
     4141 tgacagacgc gacggcgaca agcaaacatg ctgtgcgacg ctggcgatat caaaattgct
     4201 gtctgccagg tgatcgctga tgtactgaca agcctcgcgt acccgattat ccatcggtgg
     4261 atggagcgac tcgttaatcg cttccatgcg ccgcagtaac aattgctcaa gcagatttat
     4321 cgccagcagc tccgaatagc gcccttcccc ttgcccggcg ttaatgattt gcccaaacag
     4381 gtcgctgaaa tgcggctggt gcgcttcatc cgggcgaaag aaccccgtat tggcaaatat
     4441 tgacggccag ttaagccatt catgccagta ggcgcgcgga cgaaagtaaa cccactggtg
     4501 ataccattcg cgagcctccg gatgacgacc gtagtgatga atctctcctg gcgggaacag
     4561 caaaatatca cccggtcggc aaacaaattc tcgtccctga tttttcacca ccccctgacc
     4621 gcgaatggtg agattgagaa tataaccttt cattcccagc ggtcggtcga taaaaaaatc
     4681 gagataaccg ttggcctcaa tcggcgttaa acccgccacc agatgggcat taaacgagta
     4741 tcccggcagc aggggatcat tttgcgcttc agccatactt ttcatactcc cgccattcag
     4801 ag
//
