LOCUS       sequence-469256-        6878 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  over-express E. coli hk/P-dnaN (Pol III beta sliding clamp subunit, 
            N-terminal cleveable his-kinase tag).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6878)
  AUTHORS   O'Donnell Lab
  TITLE     O'Donnell Lab Batch 1
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 6878)
  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: 
            "unpublished"
COMMENT     Sequence Label: Ec dnaN-pHK/P
FEATURES             Location/Qualifiers
     source          1..6878
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(674..1465,1466..1534))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1297..1316
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1535..1639)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1707..1725
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(1752..1757,1758..1774,1775..1780)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(1955..2002)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     1998..2016
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(3171..3194)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(3201..3212)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(3249..3260)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(3273..3302)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             3315..3337
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    3352..3376
                     /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)."
     primer_bind     complement(3376..3395)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3377..3395)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3479..3498)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        3704..3781
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     complement(3801..3820)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             6175..6366
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(6360..6382)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    6468..6607
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     6523..6542
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6622..6639)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(6793..6878,1..503))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(join(6873..6878,1..14))
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
       61 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
      121 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
      181 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
      241 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
      301 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
      361 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
      421 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
      481 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
      541 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
      601 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
      661 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
      721 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
      781 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
      841 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
      901 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
      961 atagtttgcg caacgttgtt gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg
     1021 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     1081 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     1141 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     1201 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     1261 ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca catagcagaa
     1321 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     1381 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     1441 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     1501 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     1561 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     1621 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     1681 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcaag
     1741 aattctcatg tttgacagct tatcatcgat aagctttaat gcggtagttt atcacagtta
     1801 aattgctaac gcagtcaggc accgtgtatg aaatctaaca atgcgctcat cgtcatcctc
     1861 ggcaccgtca ccctggatgc tgtaggcata ggcttggtta tgccggtact gccgggcctc
     1921 ttgcgggata tccggatata gttcctcctt tcagcaaaaa acccctcaag acccgtttag
     1981 aggccccaag gggttatgct agttattgct cagcggtggc agcagccaac tcagcttcct
     2041 ttcgggcttt gttagcagcc ggatccttac agtctcattg gcatgacaac ataagccgcg
     2101 ctctggctgg ccgcatcttc aatctgcacg ctggaaaccg aatcggtcag catcatgcgg
     2161 acgttttcgc atttcagcgc gttcagaaca tccagcacat aactgacgtt gaagccgatt
     2221 tccatctccg caccgctata ggtaacgtcg aggatctctt ccgcttcttc ctgttccggg
     2281 ttgttggcgg tgattttcag ctggttttcg ctgacataaa gacgtacgcc gcggaatttc
     2341 tcgttagaga gaatcgccgc gcgagcaaac gcctgcttga gcagatcgca gccagcttcc
     2401 agatgtttgt ccgggttctt cggcagaacg cggcgataat ccgggaagcg accatccacc
     2461 agtttggagg tgaagataaa gtcgccaacg tgggcgcgaa tgttgttgct gccaatctgt
     2521 acgcgcagcg gattgtcgcc gccgtcgagc atacgcatca gttcaatcac gcctttacgc
     2581 ggtacgatca ccgaatggct tggcaaagat tgaccaattg gcattgaaca gaccgccaga
     2641 cggtggccgt cggttgccac ggtgcgcagt tcttcacctt cggtttcaaa cagcatacca
     2701 tttaagtaat agcgaacgtc ctgatgcgcc atagaaaact gggtcgcttc aatcagacgc
     2761 ttcatcgttg cctgcggcag ggtaaattcg acttcactct gccagtcatc gaggttcggg
     2821 aaatccgccg ctggcagggt agacagcgaa aaacggctac gcccggagcg taccagcatc
     2881 cgttcacctt ccagctgcac ggcaatttcc gcgccttcag gcagaccacg gcagatatca
     2941 aagaatttgc gcgccggaac ggtcgtcgct cctggctcgt gtggctgaac cagcgcaaca
     3001 cgtgccacca tttccatctc gagatcagta ccggtcagcg acaacgtacc gtcagcaacc
     3061 tgtaacagca gattaccgag aatcggtagc gtaggacgac cacctaacgg accgctcacc
     3121 tgttgtagcg gttttaataa atgctcacgt tctacggtaa atttcatatg gggcccctgg
     3181 aacagaactt ccagaatatg gcggccttca atatggccgc tgctcacgct cgcgcggcgc
     3241 agaatatgac gaccttcgat atggccgctg ctgtgatgat gatgatgatg atgatgatga
     3301 tggcccatgg tatatctcct tcttaaagtt aaacaaaatt atttctagag gggaattgtt
     3361 atccgctcac aattccccta tagtgagtcg tattaatttc gcgggatcga gatctcgatc
     3421 ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt tgctggcgcc
     3481 tatatcgccg acatcaccga tggggaagat cgggctcgcc acttcgggct catgagcgct
     3541 tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg cgccatctcc
     3601 ttgcatgcac cattccttgc ggcggcggtg ctcaacggcc tcaacctact actgggctgc
     3661 ttcctaatgc aggagtcgca taagggagag cgtcgagatc ccggacacca tcgaatggcg
     3721 caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa
     3781 tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt
     3841 ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc
     3901 ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca
     3961 gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt
     4021 cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga
     4081 acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag
     4141 tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg
     4201 cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat
     4261 tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca
     4321 gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg
     4381 ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg
     4441 gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac
     4501 tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc
     4561 cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc
     4621 atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag
     4681 cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc
     4741 cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg
     4801 cgcgttggcc gattcattaa tgcagctggc acgacagttt cccgactgga aagcgggcag
     4861 tgagcgcaac gcaattaatg taagttagct cactcattag gcaccgggat ctcgaccgat
     4921 gcccttgaga gccttcaacc cagtcagctc cttccggtgg gcgcggggca tgactatcgt
     4981 cgccgcactt atgactgtct tctttatcat gcaactcgta ggacaggtgc cggcagcgct
     5041 ctgggtcatt ttcggcgagg accgctttcg ctggagcgcg acgatgatcg gcctgtcgct
     5101 tgcggtattc ggaatcttgc acgccctcgc tcaagccttc gtcactggtc ccgccaccaa
     5161 acgtttcggc gagaagcagg ccattatcgc cggcatggcg gccgacgcgc tgggctacgt
     5221 cttgctggcg ttcgcgacgc gaggctggat ggccttcccc attatgattc ttctcgcttc
     5281 cggcggcatc gggatgcccg cgttgcaggc catgctgtcc aggcaggtag atgacgacca
     5341 tcagggacag cttcaaggat cgctcgcggc tcttaccagc ctaacttcga tcattggacc
     5401 gctgatcgtc acggcgattt atgccgcctc ggcgagcaca tggaacgggt tggcatggat
     5461 tgtaggcgcc gccctatacc ttgtctgcct ccccgcgttg cgtcgcggtg catggagccg
     5521 ggccacctcg acctgaatgg aagccggcgg cacctcgcta acggattcac cactccaaga
     5581 attggagcca atcaattctt gcggagaact gtgaatgcgc aaaccaaccc ttggcagaac
     5641 atatccatcg cgtccgccat ctccagcagc cgcacgcggc gcatctcggg cagcgttggg
     5701 tcctggccac gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg
     5761 ggttgcctta ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc
     5821 tgctgcaaaa cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg
     5881 taaagtctgg aaacgcggaa gtcagcgccc tgcaccatta tgttccggat ctgcatcgca
     5941 ggatgctgct ggctaccctg tggaacacct acatctgtat taacgaagcg ctggcattga
     6001 ccctgagtga tttttctctg gtcccgccgc atccataccg ccagttgttt accctcacaa
     6061 cgttccagta accgggcatg ttcatcatca gtaacccgta tcgtgagcat cctctctcgt
     6121 ttcatcggta tcattacccc catgaacaga aatccccctt acacggaggc atcagtgacc
     6181 aaacaggaaa aaaccgccct taacatggcc cgctttatca gaagccagac attaacgctt
     6241 ctggagaaac tcaacgagct ggacgcggat gaacaggcag acatctgtga atcgcttcac
     6301 gaccacgctg atgagcttta ccgcagctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     6361 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     6421 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     6481 cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg
     6541 tactgagagt gcaccattgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     6601 catcaggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     6661 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     6721 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     6781 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     6841 aagtcagagg tggcgaaacc cgacaggact ataaagat
//
