LOCUS       sequence-446507-        4028 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low copy cloning vector for integration of C-terminal FLAG epitope 
            tag (AmpR). SmaI restriction site immediately upstream of the FLAG 
            epitope. FLAG epitope in same orientation as lac promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4028)
  AUTHORS   Braun HG, Kanwal N, Rivera Lopez LF, Thomassin JL
  TITLE     Generation of a plasmid series for rapid sub-cloning and use in 
            various Enterobacteriaceae.
  JOURNAL   J Biosci Bioeng. 2024 Sep 6:S1389-1723(24)00253-6. doi: 
            10.1016/j.jbiosc.2024.08.006.
  PUBMED    39244484
REFERENCE   2  (bases 1 to 4028)
  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: "J Biosci 
            Bioeng. 2024 Sep 6:S1389-1723(24)00253-6. doi: 
            10.1016/j.jbiosc.2024.08.006."
COMMENT     Sequence Label: pHBJT007
FEATURES             Location/Qualifiers
     source          1..4028
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             185..1135
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     CDS             complement(join(1887..2678,2679..2747))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2510..2529
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2748..2852)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3239..3261
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(3297..3320)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3374..3390)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3374..3390)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3387..3409)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3398..3414
                     /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)."
     promoter        complement(join(3422..3428,3429..3446,3447..3452))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3467..3488
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      join(3943..4028,1..137)
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
ORIGIN
        1 aaaaaaaaca cacaaaggtc tagcggaatt tacagagggt ctagcagaat ttacaagttt
       61 tccagcaaag gtctagcaga atttacagat acccacaact caaaggaaaa ggactagtaa
      121 ttatcattga ctagcccatc tcaattggta tagtgattaa aatcacctag accaattgag
      181 atgtatgtct gaattagttg ttttcaaagc aaatgaacta gcgattagtc gctatgactt
      241 aacggagcat gaaaccaagc taattttatg ctgtgtggca ctactcaacc ccacgattga
      301 aaaccctaca aggaaagaac ggacggtatc gttcacttat aaccaatacg ctcagatgat
      361 gaacatcagt agggaaaatg cttatggtgt attagctaaa gcaaccagag agctgatgac
      421 gagaactgtg gaaatcagga atcctttggt taaaggcttt gagattttcc agtggacaaa
      481 ctatgccaag ttctcaagcg aaaaattaga attagttttt agtgaagaga tattgcctta
      541 tcttttccag ttaaaaaaat tcataaaata taatctggaa catgttaagt cttttgaaaa
      601 caaatactct atgaggattt atgagtggtt attaaaagaa ctaacacaaa agaaaactca
      661 caaggcaaat atagagatta gccttgatga atttaagttc atgttaatgc ttgaaaataa
      721 ctaccatgag tttaaaaggc ttaaccaatg ggttttgaaa ccaataagta aagatttaaa
      781 cacttacagc aatatgaaat tggtggttga taagcgaggc cgcccgactg atacgttgat
      841 tttccaagtt gaactagata gacaaatgga tctcgtaacc gaacttgaga acaaccagat
      901 aaaaatgaat ggtgacaaaa taccaacaac cattacatca gattcctacc tacataacgg
      961 actaagaaaa acactacacg atgctttaac tgcaaaaatt cagctcacca gttttgaggc
     1021 aaaatttttg agtgacatgc aaagtaagta tgatctcaat ggttcgttct catggctcac
     1081 gcaaaaacaa cgaaccacac tagagaacat actggctaaa tacggaagga tctgaggttc
     1141 ttatggctct tgtatctatc agtgaagcat caagactaac aaacaaaagt agaacaactg
     1201 ttcaccgtta catatcaaag ggaaaactgt ccatatgcac agatgaaaac ggtgtaaaaa
     1261 agatagatac atcagagctt ttacgagttt ttggtgcatt caaagctgtt caccatgaac
     1321 agatcgacaa tgtaacagat gaacagcatg taacacctaa tagaacaggt gaaaccagta
     1381 aaacaaagca actagaacat gaaattgaac acctgagaca acttgttaca gctcaacagt
     1441 cacacataga cagcctgaaa caggcgatgc tgcttatcga atcaaagctg ccgacaacac
     1501 gggagccagt gacgcctccc gtggggaaaa aatcatggca attctggaag aaatagcgct
     1561 ttcagccggc aaaccggctg aagccggatc tgcgattctg ataacaaact agcaacacca
     1621 gaacagcccg tttgcgggca gcaaaacccg tacttttgga cgttccggcg gttttttgtg
     1681 gcgagtggtg ttcgggcggt gcgcgcaaga tccattatgt taaacgggcg agtttacatc
     1741 tcaaaaccgc ccgcttaaca ccatcagaaa tcctcagcgc gattttaagc accaaccccc
     1801 cccccgtaac acccaaatcc atactgaaag tggctttgtt gaataaatca gatttcgggt
     1861 aagtctccta ggcaccaata actgccttac caatgcttaa tcagtgaggc acctatctca
     1921 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     1981 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     2041 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     2101 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     2161 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     2221 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     2281 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     2341 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     2401 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     2461 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     2521 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     2581 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     2641 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     2701 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     2761 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     2821 atttagaaaa ataaacaaat aggggttccg cgacgtctca ttttcgccaa aagttggccc
     2881 agggcttccc ggtatcaaca gggacaccag gatttattta ttctgcgaag tgatcttccg
     2941 tcacaggtat ttattcggcg caaagtgcgt cgggtgatgc tgccaactta ctgatttagt
     3001 gtatgatggt gtttttgagg tgctccagtg gcttctgttt ctatcagctg tccctcctgt
     3061 tcagctactg acggggtggt gcgtaacggc aaaagcaccg ccggacatca gcgccattcg
     3121 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     3181 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     3241 cagtcacgac gttgtaaaac gacggccgtg aattcgagct cggtacccta tcattacttg
     3301 tcgtcatcgt ctttgtagtc cccggggatc ctctagagtc gacctgcagg catgcaagct
     3361 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     3421 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     3481 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     3541 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcctcgc
     3601 taacggattc accactccgc agacccgcca taaaacgccc tgagaagccc gtgacgggct
     3661 tttcttgtat tatgggtagt ttccttgcat gaatccataa aaggcgcctg tagtgccatt
     3721 tacccccatt cactgccaga gccgtgagcg cagcgaactg aatgtcacga aaaagacagc
     3781 gactcaggtg cctgatggtc ggagacaaaa ggaatattca gcgatttgcc cgagcttgcg
     3841 agggtgctac ttaagccttt agggttttaa ggtctgtttt gtagaggagc aaacagcgtt
     3901 tgcgacatcc ttttgtaata ctgcggaact gactaaagta gtgagttata cacagggctg
     3961 ggatctattc tttttatctt tttttattct ttctttattc tataaattat aaccacttga
     4021 atataaac
//
