LOCUS       sequence-176857-        4808 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express PhoCl in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4808)
  AUTHORS   Zhang W, Lohman AW, Zhuravlova Y, Lu X, Wiens MD, Hoi H, Yaganoglu 
            S, Mohr MA, Kitova EN, Klassen JS, Pantazis P, Thompson RJ, Campbell
            RE
  TITLE     Optogenetic control with a photocleavable protein, PhoCl.
  JOURNAL   Nat Methods. 2017 Mar 13. doi: 10.1038/nmeth.4222.
  PUBMED    28288123
REFERENCE   2  (bases 1 to 4808)
  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: "Nat 
            Methods. 2017 Mar 13. doi: 10.1038/nmeth.4222."
COMMENT     Sequence Label: pBAD/HisB-PhoCl
FEATURES             Location/Qualifiers
     source          1..4808
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        15..299
                     /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     222..241
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             345..362
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             366..398
                     /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(402..410,411..425)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             441..1166
                     /codon_start=1
                     /product="photocleavable protein that separates into two 
                     fragments upon exposure to violet light (Zhang et al., 
                     2017)"
                     /label=PhoCl
                     /translation="VIPDYFKQSFPEGYSWERSMTYEDGGICIATNDITMEGDSFINKI
                     HFKGTNFPPNGPVMQKRTVGWEASTEKMYERDGVLKGDVKMKLLLKGGGHYRCDYRTTY
                     KVKQKPVKLPDYHFVDHRIEILSHDKDYNKVKLYEHAVARNSTDSMDELYKGGSGGMVS
                     KGEETITSVIKPDMKNKLRMEGNVNGHAFVIEGEGSGKPFEGIQTIDLEVKEGAPLPFA
                     YDILTTAFHYGNRVFTKYPR"
     primer_bind     complement(1223..1240)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1393..1479
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1571..1598
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1617..1708
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1709..1777,1778..2569)
                     /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(1927..1946)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2740..3328
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3229..3248
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3482..3499
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3514..3654
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3580..3599)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3740..3762
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3918..4796)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 cccgccattc agagaagaaa ccaattgtcc atattgcatc agacattgcc gtcactgcgt
       61 cttttactgg ctcttctcgc taaccaaacc ggtaaccccg cttattaaaa gcattctgta
      121 acaaagcggg accaaagcca tgacaaaaac gcgtaacaaa agtgtctata atcacggcag
      181 aaaagtccac attgattatt tgcacggcgt cacactttgc tatgccatag catttttatc
      241 cataagatta gcggatccta cctgacgctt tttatcgcaa ctctctactg tttctccata
      301 cccgtttttt gggctaacag gaggaattaa ccatgggggg ttctcatcat catcatcatc
      361 atggtatggc tagcatgact ggtggacagc aaatgggtcg ggatctgtac gacgatgacg
      421 ataaggatcc gagctcgagt gtgatccctg actacttcaa gcagagcttc cccgagggct
      481 acagctggga gcgcagcatg acctacgagg acggcggcat ctgcatcgcc accaacgaca
      541 tcacaatgga gggggacagc ttcatcaaca agatccactt caagggcacg aacttccccc
      601 ccaacggccc cgtgatgcag aagaggaccg tgggctggga ggccagcacc gagaagatgt
      661 acgagcgcga cggcgtgctg aagggcgacg tgaagatgaa gctgctgctg aagggcggcg
      721 gccactatcg ctgcgactac cgcaccacct acaaggtcaa gcagaagccc gtaaagctgc
      781 ccgactacca cttcgtggac caccgcatcg agatcctgag ccacgacaag gactacaaca
      841 aggtgaagct gtacgagcac gccgtggccc gcaactccac cgacagcatg gacgagctgt
      901 acaagggtgg cagcggtggc atggtgagca agggcgagga gaccattaca agcgtgatca
      961 agcctgacat gaagaacaag ctgcgcatgg agggcaacgt gaacggccac gccttcgtga
     1021 tcgagggcga gggcagcggc aagcccttcg agggcatcca gacgattgat ttggaggtga
     1081 aggagggcgc cccgctgccc ttcgcctacg acatcctgac caccgccttc cactacggca
     1141 accgcgtgtt caccaagtac ccacggggag gtggaggtac ctaaaagctt ggctgttttg
     1201 gcggatgaga gaagattttc agcctgatac agattaaatc agaacgcaga agcggtctga
     1261 taaaacagaa tttgcctggc ggcagtagcg cggtggtccc acctgacccc atgccgaact
     1321 cagaagtgaa acgccgtagc gccgatggta gtgtggggtc tccccatgcg agagtaggga
     1381 actgccaggc atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc
     1441 tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc cgccgggagc ggatttgaac
     1501 gttgcgaagc aacggcccgg agggtggcgg gcaggacgcc cgccataaac tgccaggcat
     1561 caaattaagc agaaggccat cctgacggat ggcctttttg cgtttctaca aactcttttg
     1621 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     1681 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     1741 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     1801 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     1861 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     1921 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
     1981 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     2041 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     2101 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     2161 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     2221 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     2281 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     2341 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     2401 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     2461 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     2521 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     2581 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     2641 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     2701 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     2761 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     2821 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     2881 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     2941 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3001 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     3061 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     3121 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     3181 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     3241 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     3301 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     3361 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     3421 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     3481 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     3541 tctgtgcggt atttcacacc gcatatggtg cactctcagt acaatctgct ctgatgccgc
     3601 atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
     3661 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     3721 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     3781 aaacgcgcga ggcagcagat caattcgcgc gcgaaggcga agcggcatgc ataatgtgcc
     3841 tgtcaaatgg acgaagcagg gattctgcaa accctatgct actccgtcaa gccgtcaatt
     3901 gtctgattcg ttaccaatta tgacaacttg acggctacat cattcacttt ttcttcacaa
     3961 ccggcacgga actcgctcgg gctggccccg gtgcattttt taaatacccg cgagaaatag
     4021 agttgatcgt caaaaccaac attgcgaccg acggtggcga taggcatccg ggtggtgctc
     4081 aaaagcagct tcgcctggct gatacgttgg tcctcgcgcc agcttaagac gctaatccct
     4141 aactgctggc ggaaaagatg tgacagacgc gacggcgaca agcaaacatg ctgtgcgacg
     4201 ctggcgatat caaaattgct gtctgccagg tgatcgctga tgtactgaca agcctcgcgt
     4261 acccgattat ccatcggtgg atggagcgac tcgttaatcg cttccatgcg ccgcagtaac
     4321 aattgctcaa gcagatttat cgccagcagc tccgaatagc gcccttcccc ttgcccggcg
     4381 ttaatgattt gcccaaacag gtcgctgaaa tgcggctggt gcgcttcatc cgggcgaaag
     4441 aaccccgtat tggcaaatat tgacggccag ttaagccatt catgccagta ggcgcgcgga
     4501 cgaaagtaaa cccactggtg ataccattcg cgagcctccg gatgacgacc gtagtgatga
     4561 atctctcctg gcgggaacag caaaatatca cccggtcggc aaacaaattc tcgtccctga
     4621 tttttcacca ccccctgacc gcgaatggtg agattgagaa tataaccttt cattcccagc
     4681 ggtcggtcga taaaaaaatc gagataaccg ttggcctcaa tcggcgttaa acccgccacc
     4741 agatgggcat taaacgagta tcccggcagc aggggatcat tttgcgcttc agccatactt
     4801 ttcatact
//
