LOCUS       sequence-335073-        8161 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Strong human ACE2 abundance from Bxb1 landing pad.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8161)
  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 8161)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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_IRES-mCherry-H2A-P2A-PuroR
FEATURES             Location/Qualifiers
     source          1..8161
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(541..1332,1333..1401))
                     /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     1164..1183
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1402..1506)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1532..1987)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1669..1690)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1881..1900
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2114..2136
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2128..2145
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2129..2145
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2152..2171
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2152..2170
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2244..2281
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     misc_feature    join(4881..5455,5456..5458,5459..5467)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(5048..5065)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     5275..5294
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     complement(5613..5631)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5881..5900
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             6611..6667
                     /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     7161..7181
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7372..7493
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7409..7428)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7463..7482
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        complement(7502..7520)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7541..7557)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7541..7557)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7554..7576)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7565..7581
                     /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(7589..7595,7596..7613,7614..7619))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7634..7655
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7772..7789)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(7943..8161,1..370))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8023..8042)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
       61 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
      121 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
      181 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
      241 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
      301 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
      361 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
      421 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
      481 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
      541 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
      601 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
      661 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
      721 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
      781 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
      841 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
      901 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
      961 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     1021 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     1081 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     1141 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     1201 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     1261 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     1321 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     1381 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     1441 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     1501 tccgcgcaca tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa
     1561 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     1621 atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac
     1681 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     1741 ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     1801 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     1861 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     1921 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     1981 ggcgcgtccc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     2041 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
     2101 acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gtaatacgac
     2161 tcactatagg gcgaattggg taccgggccc cccctcgagg tcgacgatgt aggtcacggc
     2221 attccggact cagcgctcga gccggcttgt cgacgacggc ggtctccgtc gtcaggatca
     2281 tccgcagatt ataccgcaac tacacgccac catgtccagc tcctcctggc tccttctcag
     2341 ccttgttgct gtaactgctg ctcagtccac cattgaggaa caggccaaga catttttgga
     2401 caagtttaac cacgaagccg aagacctgtt ctatcaaagt tcacttgctt cttggaatta
     2461 taacaccaat attactgaag agaatgtcca aaacatgaat aatgctgggg acaaatggtc
     2521 tgccttttta aaggaacagt ccacacttgc ccaaatgtat ccactacaag aaattcagaa
     2581 tctcacagtc aagcttcagc tgcaggctct tcagcaaaat gggtcttcag tgctctcaga
     2641 agacaagagc aaacggttga acacaattct aaatacaatg agcaccatct acagtactgg
     2701 aaaagtttgt aacccagata atccacaaga atgcttatta cttgaaccag gtttgaatga
     2761 aataatggca aacagtttag actacaatga gaggctctgg gcttgggaaa gctggagatc
     2821 tgaggtcggc aagcagctga ggccattata tgaagagtat gtggtcttga aaaatgagat
     2881 ggcaagagca aatcattatg aggactatgg ggattattgg agaggagact atgaagtaaa
     2941 tggggtagat ggctatgact acagccgcgg ccagttgatt gaagatgtgg aacatacctt
     3001 tgaagagatt aaaccattat atgaacatct tcatgcctat gtgagggcaa agttgatgaa
     3061 tgcctatcct tcctatatca gtccaattgg atgcctccct gctcatttgc ttggtgatat
     3121 gtggggtaga ttttggacaa atctgtactc tttgacagtt ccctttggac agaaaccaaa
     3181 catagatgtt actgatgcaa tggtggacca ggcctgggat gcacagagaa tattcaagga
     3241 ggccgagaag ttctttgtat ctgttggtct tcctaatatg actcaaggat tctgggaaaa
     3301 ttccatgcta acggacccag gaaatgttca gaaagcagtc tgccatccca cagcttggga
     3361 cctggggaag ggcgacttca ggatccttat gtgcacaaag gtgacaatgg acgacttcct
     3421 gacagctcat catgagatgg ggcatatcca gtatgatatg gcatatgctg cacaaccttt
     3481 tctgctaaga aatggagcta atgaaggatt ccatgaagct gttggggaaa tcatgtcact
     3541 ttctgcagcc acacctaagc atttaaaatc cattggtctt ctgtcacccg attttcaaga
     3601 agacaatgaa acagaaataa acttcctgct caaacaagca ctcacgattg ttgggactct
     3661 gccatttact tacatgttag agaagtggag gtggatggtc tttaaagggg aaattcccaa
     3721 agaccagtgg atgaaaaagt ggtgggagat gaagcgagag atagttgggg tggtggaacc
     3781 tgtgccccat gatgaaacat actgtgaccc cgcatctctg ttccatgttt ctaatgatta
     3841 ctcattcatt cgatattaca caaggaccct ttaccaattc cagtttcaag aagcactttg
     3901 tcaagcagct aaacatgaag gccctctgca caaatgtgac atctcaaact ctacagaagc
     3961 tggacagaaa ctgttcaata tgctgaggct tggaaaatca gaaccctgga ccctagcatt
     4021 ggaaaatgtt gtaggagcaa agaacatgaa tgtaaggcca ctgctcaact actttgagcc
     4081 cttatttacc tggctgaaag accagaacaa gaattctttt gtgggatgga gtaccgactg
     4141 gagtccatat gcagaccaaa gcatcaaagt gaggataagc ctaaaatcag ctcttggaga
     4201 taaagcatat gaatggaacg acaatgaaat gtacctgttc cgatcatctg ttgcatatgc
     4261 tatgaggcag tactttttaa aagtaaaaaa tcagatgatt ctttttgggg aggaggatgt
     4321 gcgagtggct aatttgaaac caagaatctc ctttaatttc tttgtcactg cacctaaaaa
     4381 tgtgtctgat atcattccta gaactgaagt tgaaaaggcc atcaggatgt cccggagccg
     4441 tatcaatgat gctttccgtc tgaatgacaa cagcctagag tttctgggga tacagccaac
     4501 acttggacct cctaaccagc cccctgtttc catatggctg attgtttttg gagttgtgat
     4561 gggagtgata gtggttggca ttgtcatcct gatcttcact gggatcagag atcggaagaa
     4621 gaaaaataaa gcaagaagtg gagaaaatcc ttatgcctcc atcgatatta gcaaaggaga
     4681 aaataatcca ggattccaaa acactgatga tgttcagacc tcctttggta ccgagacctc
     4741 ccaggtggcg cccgcttagt aacttaagaa ttcaccggtc tgacctcctt ctccctctct
     4801 tcaggtctgc agtcgacggt accgcgggcc cgggatccac cggatctaga taactgatca
     4861 ctgtggaatc agagagactg cccctctccc tctccccccc ctaacgttac tggccgaagc
     4921 cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct
     4981 tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt
     5041 ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct
     5101 ctggaagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc
     5161 ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag
     5221 gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc
     5281 tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga
     5341 tctgatctgg ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt
     5401 ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataatatggc
     5461 cacaaccatg gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg
     5521 cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg
     5581 cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc
     5641 cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt
     5701 gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg
     5761 ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct
     5821 gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg
     5881 ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga
     5941 ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta
     6001 cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta
     6061 caacgtcaac atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca
     6121 gtacgaacgc gccgagggcc gccactccac cggcggctac tcagatctcg agctcaagct
     6181 tcgaattctg atgtcgggac gcggcaagca gggaggcaaa gctcgcgcca aagccaagac
     6241 ccgctcttct cgtgccggtc tccagttccc cgtgggccga gtgcaccgac tgctccgcaa
     6301 gggcaactat gctgagcggg tcggggccgg cgcgccggtg tacctggcgg cggtgctgga
     6361 gtacctgact gccgagatcc tggagctggc gggcaacgcc gcccgcgaca acaagaagac
     6421 ccgcattatc ccgcgccact tgcagctggc catccgcaac gacgaggagc tcaacaagct
     6481 gctgggcaaa gtaaccatcg ctcagggtgg tgtcctgccc aacatccagg ctgtgctact
     6541 gcccaagaag accgagagtc accacaaggc caaaggcaaa cgggatccac cggatctaga
     6601 taacagcgga gctactaact tcagcctgct gaagcaggct ggagacgtgg aggagaaccc
     6661 tggacctacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc
     6721 cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc
     6781 ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct
     6841 cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc
     6901 ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag
     6961 cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa
     7021 ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct
     7081 gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt
     7141 cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac
     7201 cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agtaataggg
     7261 atcctgatca taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc
     7321 ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt
     7381 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     7441 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaggagatc
     7501 ccctttagtg agggttaatt tcgagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     7561 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     7621 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     7681 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     7741 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cactcggtcg
     7801 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     7861 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     7921 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     7981 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     8041 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     8101 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     8161 g
//
