LOCUS       sequence-335070-        6106 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ectodomain deleted negative control ACE2 from Bxb1 landing pad.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6106)
  AUTHORS   Shukla N, Roelle SM, Suzart VG, Bruchez AM, Matreyek KA
  TITLE     Mutants of human ACE2 differentially promote SARS-CoV and SARS-CoV-2
            spike mediated infection.
  JOURNAL   PLoS Pathog. 2021 Jul 16;17(7):e1009715. doi: 
            10.1371/journal.ppat.1009715. eCollection 2021 Jul.
  PUBMED    34270613
REFERENCE   2  (bases 1 to 6106)
  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: "PLoS 
            Pathog."; date: "2021-07-16"; volume: "17(7):e1009715. doi"; pages: 
            " 10.1371/journal.ppat.1009715. eCollection 2021 Jul"
COMMENT     Sequence Label: AttB_ACE2[dEcto]_IRES-mCherry-H2A-P2A-PuroR
FEATURES             Location/Qualifiers
     source          1..6106
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     163..182
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     416..433
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    550..571
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(586..591,592..609,610..616)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    624..640
                     /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     629..651
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     648..664
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     648..664
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        685..703
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     polyA_signal    712..833
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(723..742)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     777..796
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1024..1044)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             complement(1538..1594)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     primer_bind     complement(2305..2324)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2574..2592
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     misc_feature    join(2738..2746,2747..2749,2750..3324)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2911..2930)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     3140..3157
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     protein_bind    3869..3906
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     primer_bind     complement(3979..3998)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3980..3998)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4005..4022)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4005..4021)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4014..4036)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4163..4618
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4250..4269)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4460..4481
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4644..4748
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4749..4817,4818..5609)
                     /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     complement(4967..4986)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(5780..6106,1..262)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
       61 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
      121 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
      181 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
      241 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
      301 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
      361 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agtgcagcga
      421 gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
      481 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
      541 caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct
      601 tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta
      661 tgaccatgat tacgccaagc tcgaaattaa ccctcactaa aggggatctc ctaagataca
      721 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
      781 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca
      841 acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca
      901 agtaaaacct ctacaaatgt ggtatggctg attatgatca ggatccctat tacttgcggg
      961 tcatgcacca ggtgcgcggt ccttcgggca cctcgacgtc ggcggtgacg gtgaagccga
     1021 gccgctcgta gaaggggagg ttgcggggcg cggaggtctc caggaaggcg ggcaccccgg
     1081 cgcgctcggc cgcctccact ccggggagca cgacggcgct gcccagaccc ttgccctggt
     1141 ggtcgggcga gacgccgacg gtggccagga accacgcggg ctccttgggc cggtgcggcg
     1201 ccaggaggcc ttccatctgt tgctgcgcgg ccagccggga accgctcaac tcggccatgc
     1261 gcgggccgat ctcggcgaac accgcccccg cttcgacgct ctccggcgtg gtccagaccg
     1321 ccaccgcggc gccgtcgtcc gcgacccaca ccttgccgat gtcgagcccg acgcgcgtga
     1381 ggaagagttc ttgcagctcg gtgacccgct cgatgtggcg gtccggatcg acggtgtggc
     1441 gcgtggcggg gtagtcggcg aacgcggcgg cgagggtgcg tacggccctg gggacgtcgt
     1501 cgcgggtggc gaggcgcacc gtgggcttgt actcggtagg tccagggttc tcctccacgt
     1561 ctccagcctg cttcagcagg ctgaagttag tagctccgct gttatctaga tccggtggat
     1621 cccgtttgcc tttggccttg tggtgactct cggtcttctt gggcagtagc acagcctgga
     1681 tgttgggcag gacaccaccc tgagcgatgg ttactttgcc cagcagcttg ttgagctcct
     1741 cgtcgttgcg gatggccagc tgcaagtggc gcgggataat gcgggtcttc ttgttgtcgc
     1801 gggcggcgtt gcccgccagc tccaggatct cggcagtcag gtactccagc accgccgcca
     1861 ggtacaccgg cgcgccggcc ccgacccgct cagcatagtt gcccttgcgg agcagtcggt
     1921 gcactcggcc cacggggaac tggagaccgg cacgagaaga gcgggtcttg gctttggcgc
     1981 gagctttgcc tccctgcttg ccgcgtcccg acatcagaat tcgaagcttg agctcgagat
     2041 ctgagtagcc gccggtggag tggcggccct cggcgcgttc gtactgttcc acgatggtgt
     2101 agtcctcgtt gtgggaggtg atgtccaact tgatgttgac gttgtaggcg ccgggcagct
     2161 gcacgggctt cttggccttg taggtggtct tgacctcagc gtcgtagtgg ccgccgtcct
     2221 tcagcttcag cctctgcttg atctcgccct tcagggcgcc gtcctcgggg tacatccgct
     2281 cggaggaggc ctcccagccc atggtcttct tctgcattac ggggccgtcg gaggggaagt
     2341 tggtgccgcg cagcttcacc ttgtagatga actcgccgtc ctgcagggag gagtcctggg
     2401 tcacggtcac cacgccgccg tcctcgaagt tcatcacgcg ctcccacttg aagccctcgg
     2461 ggaaggacag cttcaagtag tcggggatgt cggcggggtg cttcacgtag gccttggagc
     2521 cgtacatgaa ctgaggggac aggatgtccc aggcgaaggg cagggggcca cccttggtca
     2581 ccttcagctt ggcggtctgg gtgccctcgt aggggcggcc ctcgccctcg ccctcgatct
     2641 cgaactcgtg gccgttcacg gagccctcca tgtgcacctt gaagcgcatg aactccttga
     2701 tgatggccat gttatcctcc tcgcccttgc tcaccatggt tgtggccata ttatcatcgt
     2761 gtttttcaaa ggaaaaccac gtccccgtgg ttcggggggc ctagacgttt ttttaacctc
     2821 gactaaacac atgtaaagca tgtgcaccga ggccccagat cagatcccat acaatggggt
     2881 accttctggg catccttcag ccccttgttg aatacgcttg aggagagcca tttgactctt
     2941 tccacaacta tccaactcac aacgtggcac tggggttgtg ccgcctttgc aggtgtatct
     3001 tatacacgtg gcttttggcc gcagaggcac ctgtcgccag gtggggggtt ccgctgcctg
     3061 caaagggtcg ctacagacgt tgtttgtctt caagaagctt ccagaggaac tgcttccttc
     3121 acgacattca acagaccttg cattcctttg gcgagagggg aaagacccct aggaatgctc
     3181 gtcaagaaga cagggccagg tttccgggcc ctcacattgc caaaagacgg caatatggtg
     3241 gaaaataaca tatagacaaa cgcacaccgg ccttattcca agcggcttcg gccagtaacg
     3301 ttaggggggg gagagggaga ggggcagtct ctctgattcc acagtgatca gttatctaga
     3361 tccggtggat cccgggcccg cggtaccgtc gactgcagac ctgaagagag ggagaaggag
     3421 gtcagaccgg tgaattctta agttactaag cgggcgccac ctgggaggtc tcggtaccaa
     3481 aggaggtctg aacatcatca gtgttttgga atcctggatt attttctcct ttgctaatat
     3541 cgatggaggc ataaggattt tctccacttc ttgctttatt tttcttcttc cgatctctga
     3601 tcccagtgaa gatcaggatg acaatgccaa ccactatcac tcccatcaca actccaaaaa
     3661 caatcagcca tatggaaaca gggggctggt taggaggtcc aagtgttggc tgtatcccca
     3721 gaaactctag gctgttgtca ttcagacgga aagcatcatt gatacggctt tcctcaatgg
     3781 tggactgagc agcagttaca gcaacaaggc tgagaaggag ccaggaggag ctggacatgg
     3841 tggcgtgtag ttgcggtata atctgcggat gatcctgacg acggagaccg ccgtcgtcga
     3901 caagccggct cgagcgctga gtccggaatg ccgtgaccta catcgtcgac ctcgaggggg
     3961 ggcccggtac ccaattcgcc ctatagtgag tcgtattaca attcactggc cgtcgtttta
     4021 caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc
     4081 cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg
     4141 cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     4201 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     4261 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     4321 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     4381 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     4441 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     4501 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     4561 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag
     4621 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     4681 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     4741 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     4801 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     4861 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     4921 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     4981 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     5041 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     5101 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     5161 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     5221 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     5281 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     5341 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     5401 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     5461 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     5521 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     5581 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     5641 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     5701 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     5761 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     5821 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     5881 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     5941 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     6001 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     6061 gacgatagtt accggataag gcgcagcggt cgggctgaac gggggg
//
