LOCUS       sequence-389738-        7082 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express NtA-Fs1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7082)
  TITLE     Jeff Miner Lab Plasmids
REFERENCE   2  (bases 1 to 7082)
  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"
COMMENT     Sequence Label: pCI_E01_NtA-Fs1-Myc-His-WPRE
FEATURES             Location/Qualifiers
     source          1..7082
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(177..968,969..1037))
                     /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     800..819
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1038..1142)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    1298..1346
                     /note="synthetic polyadenylation signal"
     CDS             complement(1370..2407)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1383..1391
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(2438..2795)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2452..2587
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        2604..2795
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     2752..2772
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    3040..3161
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3077..3096)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3131..3150
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3198..3216
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     misc_feature    3304..3892
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3410..3421
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3819..3839
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3929..3946)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(3974..4003)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(4013..4042)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(4052..4081)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(4091..4120)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(4145..4165)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     regulatory      4900..4909
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(5009..5028)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5010..5028)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5070..5089)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          5072..5204
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        complement(5340..5543)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5369..5389)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5544..5923
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     misc_feature    6114..6397
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     rep_origin      join(6513..7082,1..19)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7002..7021
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
       61 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
      121 taccgccttt gagtgagctg ataccggttg ggcgtcgctt ggtcggtcat ttcgaattac
      181 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
      241 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
      301 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
      361 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
      421 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
      481 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
      541 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
      601 agctccttcg gtcctccgat agttgtcaga agtaagttgg ccgcagtgtt atcactcatg
      661 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
      721 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
      781 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttgaa agtgctcatc
      841 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
      901 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
      961 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     1021 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     1081 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     1141 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattacgta gaaagccagt
     1201 ccgcagaaac ggtgctgacc ccggatgaat gtcagctact gggctatctg gacaagggaa
     1261 aacgcaagcg caaagaggat ccttatcgct atcgattcac acaaaaaacc aacacacaga
     1321 tgtaatgaaa ataaagatat tttattgcgg ccatcgtgat ggctagagtc taaactcgac
     1381 ctacctcctt agcgcgcggt cgtgtactgg gccggcggtt gccgctgtcg gccagcacct
     1441 cgacgcagcc gtcggtccat acggctgcgc tccggcgagc gatttgtgtg cgaccgacgg
     1501 tgccggcccc gctgcggacg atggcatcgc agcggccttg agcccaggca gcatcgtcga
     1561 agttgccgtc cacgaggctc tggtagagct gatccaggcc gatgcgcagc atgtaggctc
     1621 gcagacgagg gctgccggcc agctcgggat gccggcgctc gaagtagcga gtctgctgct
     1681 ccatgcaagc cagccagggc cgccagaaga agatgttggc cacctcgtac tgactgtccc
     1741 cgaacatagc ttcggaccag tcgattacgg cggtgatgcg gccgttgtcg gtcaggacgt
     1801 tgttgctgcc gaagtcggca tggacgaggt ggcgcacctc gggacagtct tcggcccaca
     1861 gcatcagttc gtccagggct tgagctacgc tggcggacac ggtgtcgtcc atcacggtct
     1921 gccagtggta gacatgggga tcagcaatgg cgcaaatgaa atcccgccag gttgtgtact
     1981 ggccgatgcc ttggggcccg aatgggccga agccgctggt ttgactgagg tcggcggcgg
     2041 caatagcatc catagcttcg gcgacaggct gtaacacagc aggcagctct gtttcgggga
     2101 ggtcttggag agtaacgcct tgggcgcgtc tactgatgca gtatgtcagg ctctcgctga
     2161 actcgccgat gtccaacact tcggggatgg gtagtgcagc gctggcgaag tggcggtaca
     2221 cgtagcggtc tttgtagaag ccatcagcgc agctattcac ccgcagtaca tagccgcgtc
     2281 cgccgacatc gaagctgaag gctcggctct cttcgccctc cgacaactgc atcaggtcgc
     2341 tcacactgtc gaacttctcg atgagaaatt tttcaacgct ggtagcggtg agttcgggct
     2401 tcttcatggt ggcgctagtg tcagaagaat cgagcttttt gcaaaagcct aggcctccaa
     2461 aaaagcctcc tcactacttc tggaatagct cagaggccga ggcggcctcg gcctctgcat
     2521 aaataaaaaa aattagtcag ccatggggcg gagaatgggc ggaactgggc ggagttaggg
     2581 gcgggatggg cggagttagg ggcgggacta tggttgctga ctaattgaga tgcatgcttt
     2641 gcatacttct gcctgctggg gagcctgggg actttccaca cctggttgct gactaattga
     2701 gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca caccctaact
     2761 gacacacatt ccacagctgg ttctttccgc ctcagaaggt acctaaccaa gttcctcttt
     2821 cagaggttat ttcaggccat ggtgctgcgc agatccgcgt atgcggtgtg aaataccgca
     2881 cagatgcgta aggagaaaat accgcatcag gaaattgtaa gcgttaatat tttgttaaaa
     2941 ttcgatttta ccacatttgt agaggtttta cttgcttaaa aaaacctccc acatctcccc
     3001 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     3061 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3121 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg ctcgaagcgg
     3181 ccgccccaag gggttatgct agttattgct cagcggtggc agcagccaac tcagcttcct
     3241 ttcgggcttt gttagcagcc ggatcagctt gcatgcccta gagtcgcggc ccaaggaacc
     3301 tgggcgggga ggcggcccaa agggagatcc gactcgtctg agggcgaagg cgaagacgcg
     3361 gaagaggccg cagagccggc agcaggccgc gggaaggaag gtccgctgga ttgagggccg
     3421 aagggacgta gcagaaggac gtcccgcgca gaatccaggt ggcaacacag gcgagcagcc
     3481 atggaaagga cgtcagcttc cccgacaaca ccacggaatt gtcagtgccc aacagccgag
     3541 cccctgtcca gcagcgggca aggcaggcgg cgatgagttc cgccgtggca atagggaggg
     3601 ggaaagcgaa agtcccggaa aggagctgac aggtggtggc aatgccccaa ccagtggggg
     3661 ttgcgtcagc aaacacagtg cacaccacgc cacgttgcct gacaacgggc cacaactcct
     3721 cataaagaga cagcaaccag gatttataca aggaggagaa aatgaaagcc atacgggaag
     3781 caatagcatg atacaaaggc attaaagcag cgtatccaca tagcgtaaaa ggagcaacat
     3841 agttaagaat accagtcaat ctttcacaaa ttttgtaatc cagaggttga ttgtcgatga
     3901 atttctagaa ttcagctgat atcccctagt ggtgatggtg atgatggagg tcgcccaagc
     3961 tctccatttc attcaagtcc tcttcagaaa tgagcttttg ctccatttca ttcaagtcct
     4021 cttcagaaat gagcttttgc tccatttcat tcaagtcctc ttcagaaatg agcttttgct
     4081 ccatttcatt caagtcctct tcagaaatga gcttttgctc catagcttta ccacttccat
     4141 cgatgctctg aaagtacaga tcctcggacc cacatggccc ttggcggagt aggcggatgc
     4201 gccgttgctg gttgcactgt gcacgctgta gctcacactc gttgctatag gtggaagcat
     4261 ctgagccaca cacaggagct accatagcag ggcagacatt cttcttgcac acacaggagg
     4321 cccggcctgg atcctcaaca ctaggttcac acacagcacc aaagccacat aacattcccc
     4381 tgcacacatc aggaggcata gaaggcgctg ctgtgaagtg gatcccaggt ttgtcttcta
     4441 cacagaactc cacctcttcc aggttccgca aagtgatccg cataaggctg gagttgagca
     4501 tcaactcatt cttgtgggct ggccacaggt agggtggtgc tgggttcaca aagaagattc
     4561 tagtgtcccc ggtggacacc tggttgtcgc agatgagggg gtctccaaag ccaccaatta
     4621 ccaccttgtt gccaccatcc agcaggctct cttgggccac cacatctttg cctttcaagt
     4681 accgccacac tcgaaccttg caggagtagg tgtgctgcac cgggtccacg ttgaggatct
     4741 cctccactgt gccggtgagc accacgttcg cctcctcctc gcgccgctcc aaggcgcgct
     4801 ccgggcacgt cccgctggcg cccggcacgg ctggcgcggc cgccgccagg agcagcagca
     4861 gcggcagcag cggggccggg aagctcagcc gcgggcggac catggtggcg gctggtgggt
     4921 ggtgatgtag ataggtggtt agtggaaaga gtgattgaat gttattaggt gttggggaga
     4981 taggtgagtt cggcgatcgc tttgctagcc ctatagtgag tcgtattaat actctagcct
     5041 taagagctgt aattgaactg ggagtggaca cctgtggaga gaaaggcaaa gtggatgtca
     5101 gtaagaccaa taggtgccta tcagaaacgc aagagtcttc tctgtctcga caagcccagt
     5161 ttctattggt ctccttaaac ctgtcttgta accttgatac ttacctgccc agtgcctcac
     5221 gaccaacttc tgcagcttaa gttcgagact gttgtgtcag aagcactgac tgcgttagca
     5281 atttaactgt gataaactac cgcaataaag cttctagtga tctgacggtt cactaaacca
     5341 gctctgctta tatagacctc ccaccgtaca cgcctaccgc ccatttgcgt caacggggcg
     5401 gggttattac gacattttgg aaagtcccgt tgattttggt gccaaaacaa actcccattg
     5461 acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa accgctatcc acgcccattg
     5521 gtgtactgcc aaaaccgcat caccatggta atagcgatga ctaatacgta gatgtactgc
     5581 caagtaggaa agtcccgtaa ggtcatgtac tgggcataat gccaggcggg ccatttaccg
     5641 tcattgacgt caataggggg cggacttggc atatgataca cttgatgtac tgccaagtgg
     5701 gcagtttacc gtaaatactc cacccattga cgtcaatgga aagtccctat tggcgttact
     5761 atgggaacat acgtcattat tgacgtcaat gggcgggggt cgttgggcgg tcagccaggc
     5821 gggccattta ccgtaagtta tgtaacgcgg aactccatat atgggctatg aactaatgac
     5881 cccgtaattg attactatta ataactagtc aataatcaat gccaacatgg cggtcatatt
     5941 ggacatgagc caatataaat gtacatatta tgatatagat acaacgtatg caatggccaa
     6001 tagccaatat tgatttatgc tatataacca atgaataata tggctaatgg ccaatattga
     6061 agatctcgat cctctacgcc ggacgcatcg tggccggagt actgcggttt atctcgatgg
     6121 ctacgagggc agacagtaag tggatttacc ataatccctt aattgtacgc accgctaaaa
     6181 cgcgttcagc gcgatcacgg cagcagacag gtaaaaatgg caacaaacca cccgaaaaac
     6241 tgccgcaatc gcgcctgata aattttaacc gtatgaatac ctatgcaacc agagggtaca
     6301 ggccacatta cccccactta atccactgaa gctgccattt ttcatggttt caccatccca
     6361 gcgaagggcc atccagcgtg cgttcctgta tttccggact ataaaaggat ctaggtgaag
     6421 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     6481 tcagaccccg tagaaaagat caaaggatct tcttgaaatc ctttttttct gcgcgtaatc
     6541 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     6601 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     6661 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     6721 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     6781 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     6841 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     6901 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     6961 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     7021 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     7081 gg
//
