LOCUS       sequence-444699-        4804 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low copy cloning vector for integration of C-terminal FLAG epitope 
            tag (SmR). SmaI restriction site immediately upstream of the FLAG 
            epitope. FLAG epitope in opposite orientation of lac promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4804)
  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 4804)
  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: pHBJT014
FEATURES             Location/Qualifiers
     source          1..4804
                     /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"
     promoter        complement(3526..3628)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     4015..4037
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4029..4046
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4030..4046
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4091..4114
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(4150..4166)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4150..4166)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4163..4185)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4174..4190
                     /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(4198..4204,4205..4222,4223..4228))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4243..4264
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      join(4719..4804,1..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 ccccgtaaca cccaaatcca tactgaaagt ggctttgttg aataaatcag atttcgggta
     1861 agtctcctag gcaccaataa ctgccctagt atgacgtctg tcgcacctgc ttgatcgcgg
     1921 ccgcgatagc tagatcgcgt tgctcctctt ctccatccgc gttccaagct gcggaaaggc
     1981 acccataagc gtacgcctgg tcgagcaggc gacgcggatc gacgtccagc gcacgagaga
     2041 atgcgtccgc catctgtgca atgcgtctag gatcgagaca aaggtcgtct ctgtcagccg
     2101 gatcgtagaa catattggcg gcgccaaagc ccacttcacc gaccagaccg acgggatcta
     2161 tcaccagcca gccgcgactg gagaacatga tgttttcatg atgcagatcg ccatgtagcc
     2221 cacgcagttc cgaggcattg ctcatcattt gatcggctat aatcgccgcg tggacgtagt
     2281 cagtttgaca acctgcgttt tgatcatcgc gcgcccgctg aaacaaagct gcaaagcgat
     2341 cccggatcgg gagaagggca gaaggcaggg gttcctcaga tgcggcatac agcttcgcca
     2401 ttagttccgc tgcaatttcg gtcgcctggt agtcgccgtg ctcggcaacg atgtgagaga
     2461 gcattcgctc cccggcatat tcgagcaaca tcagattgtt ctcacgaccg agcaaccgga
     2521 ctgctcccct cccattgcgc cataccagat agtcggcccc gcgcagttca tcagcaatgt
     2581 cttctatagg tttcaatccc ttgacgattg caggagtccc gtctggcaat gaaactttcc
     2641 aaacgaggct ggaaaaggtg tccgcaatga gaacaggttg cgaaacgtgc caatgagcag
     2701 gaaaaacagg cggcatgaac atcaacccca agtcagaggg tccaatcgca gatagaaggc
     2761 aaggcgttcg cggtcggggg cttcgatccc caatacattg aataggacag cgaaggcgcg
     2821 ctctgcttca tctggcgctg cccagttctc ttcggcgtta gcaatcatga gtgccaaatc
     2881 ggcatagcga tctgctgttc cgagccgccc aaggtcgatc agacccgtgc attgaagagt
     2941 tttagggtcc accatgaagt tcggcatgca gggatcacca tggcaaacaa ccatatcggt
     3001 gcgctcttgg tcgagccgca ccggtagctc tcgttcgaca cgagccaaaa gatcgagctg
     3061 cggcgtactc ttgtcctcgt ccggtaagaa gtcgggattg acggcattgc gggacaccac
     3121 atcaacggcg cgtccgaaca ttcgcgacag cctgcgctca aacggacatt gatcaaccga
     3181 taggctgtga acagcgccaa gttgctgccc cattgacggc cacgctttga gcaaatccgc
     3241 tccagacaga tcagccgccg gtactcccgg aattgccgtt atcaccaagc atgcaccctc
     3301 ctgttcctcc tgccagttga tgacctcggg gcaagccaca cctcgacctt tgagccaaat
     3361 gaggcggtca cgctctccag cgagctcacc gcggcgggaa gcaggtgcga ttttcgcgaa
     3421 ggcatgcccg tcaccacgtc gaaaaacaaa atcaccagat tctccgcctc tgacaggcaa
     3481 ccagtcagaa tgcgattcac caaaaaaaat attagttcga ttcaatttag cttccttagc
     3541 tcctgaaaat ctcgataact caaaaaatac gcccggtagt gatcttattt cattatggtg
     3601 aaagttggaa cctcttacgt gccgatcaac gtctcatttt cgccaaaagt tggcccaggg
     3661 cttcccggta tcaacaggga caccaggatt tatttattct gcgaagtgat cttccgtcac
     3721 aggtatttat tcggcgcaaa gtgcgtcggg tgatgctgcc aacttactga tttagtgtat
     3781 gatggtgttt ttgaggtgct ccagtggctt ctgtttctat cagctgtccc tcctgttcag
     3841 ctactgacgg ggtggtgcgt aacggcaaaa gcaccgccgg acatcagcgc cattcgccat
     3901 tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc
     3961 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     4021 cacgacgttg taaaacgacg gccagtgcca agcttgcatg cctgcaggtc gactctagag
     4081 gatccccggg gactacaaag acgatgacga caagtaatga tagggtaccg agctcgaatt
     4141 cgtaatcatg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     4201 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     4261 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     4321 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cctcgctaac
     4381 ggattcacca ctccgcagac ccgccataaa acgccctgag aagcccgtga cgggcttttc
     4441 ttgtattatg ggtagtttcc ttgcatgaat ccataaaagg cgcctgtagt gccatttacc
     4501 cccattcact gccagagccg tgagcgcagc gaactgaatg tcacgaaaaa gacagcgact
     4561 caggtgcctg atggtcggag acaaaaggaa tattcagcga tttgcccgag cttgcgaggg
     4621 tgctacttaa gcctttaggg ttttaaggtc tgttttgtag aggagcaaac agcgtttgcg
     4681 acatcctttt gtaatactgc ggaactgact aaagtagtga gttatacaca gggctgggat
     4741 ctattctttt tatctttttt tattctttct ttattctata aattataacc acttgaatat
     4801 aaac
//
