LOCUS       sequence-444672-        3833 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low copy cloning vector for integration of C-terminal FLAG epitope 
            tag (ChlR). 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 3833)
  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 3833)
  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: pHBJT005
FEATURES             Location/Qualifiers
     source          1..3833
                     /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(1895..2554)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2469..2488
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2555..2657)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     3044..3066
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(3102..3125)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3179..3195)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3179..3195)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3192..3214)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3203..3219
                     /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(3227..3233,3234..3251,3252..3257))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3272..3293
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      join(3748..3833,1..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 ctgccttaaa aaaattacgc cccgccctgc cactcatcgc
     1921 agtactgttg taattcatta agcattctgc cgacatggaa gccatcacaa acggcatgat
     1981 gaacctgaat cgccagcggc atcagcacct tgtcgccttg cgtataatat ttgcccatgg
     2041 tgaaaacggg ggcgaagaag ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac
     2101 tcacccaggg attggctgag acgaaaaaca tattctcaat aaacccttta gggaaatagg
     2161 ccaggttttc accgtaacac gccacatctt gcgaatatat gtgtagaaac tgccggaaat
     2221 cgtcgtggta ttcactccag agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt
     2281 aacaagggtg aacactatcc catatcacca gctcaccgtc tttcattgcc atacgaaatt
     2341 ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa aacttgtgct
     2401 tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc tggttatagg
     2461 tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat
     2521 caacggtggt atatccagtg atttttttct ccattttagc ttccttagct cctgaaaatc
     2581 tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac
     2641 ctcttacgtg ccgatcaacg tctcattttc gccaaaagtt ggcccagggc ttcccggtat
     2701 caacagggac accaggattt atttattctg cgaagtgatc ttccgtcaca ggtatttatt
     2761 cggcgcaaag tgcgtcgggt gatgctgcca acttactgat ttagtgtatg atggtgtttt
     2821 tgaggtgctc cagtggcttc tgtttctatc agctgtccct cctgttcagc tactgacggg
     2881 gtggtgcgta acggcaaaag caccgccgga catcagcgcc attcgccatt caggctgcgc
     2941 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg
     3001 ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt
     3061 aaaacgacgg ccgtgaattc gagctcggta ccctatcatt acttgtcgtc atcgtctttg
     3121 tagtccccgg ggatcctcta gagtcgacct gcaggcatgc aagcttggcg taatcatggt
     3181 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     3241 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     3301 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     3361 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ctcgctaacg gattcaccac
     3421 tccgcagacc cgccataaaa cgccctgaga agcccgtgac gggcttttct tgtattatgg
     3481 gtagtttcct tgcatgaatc cataaaaggc gcctgtagtg ccatttaccc ccattcactg
     3541 ccagagccgt gagcgcagcg aactgaatgt cacgaaaaag acagcgactc aggtgcctga
     3601 tggtcggaga caaaaggaat attcagcgat ttgcccgagc ttgcgagggt gctacttaag
     3661 cctttagggt tttaaggtct gttttgtaga ggagcaaaca gcgtttgcga catccttttg
     3721 taatactgcg gaactgacta aagtagtgag ttatacacag ggctgggatc tattcttttt
     3781 atcttttttt attctttctt tattctataa attataacca cttgaatata aac
//
