LOCUS       sequence-313826-        7706 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminally GFP-tagged and C-terminally Flag-tagged 
            P5-fusion chimera of Pf-AMA1 and EBA175 (residues 1284-1462) from 
            pcDNA3.1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7706)
  AUTHORS   Gandhi S, Baker RP, Cho S, Stanchev S, Strisovsky K, Urban S
  TITLE     Designed Parasite-Selective Rhomboid Inhibitors Block Invasion and 
            Clear Blood-Stage Malaria.
  JOURNAL   Cell Chem Biol. 2020 Aug 31. pii: S2451-9456(20)30333-0. doi: 
            10.1016/j.chembiol.2020.08.011.
  PUBMED    32888502
REFERENCE   2  (bases 1 to 7706)
  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: "Cell Chem 
            Biol. 2020 Aug 31. pii: S2451-9456(20)30333-0. doi: 
            10.1016/j.chembiol.2020.08.011."
COMMENT     Sequence Label: pcDNA-GFP-AMA1/EBA175(P5 fusion)-Flag
FEATURES             Location/Qualifiers
     source          1..7706
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1016..1018,1019..1021,1022..1732)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1016..1018,1019..1021,1022..1732)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1061..1082)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1322..1341)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1669..1690
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3227..3250
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3297..3314)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3303..3527
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3573..4001
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3660..3679)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3870..3891
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4010..4030)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4015..4344
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4195..4330
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4257..4276
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4411..5205
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4465..4484)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5075..5094
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5379..5500
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5416..5435)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5470..5489
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5549..5565)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5549..5565)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5562..5584)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5573..5589
                     /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(5597..5603,5604..5621,5622..5627))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5642..5663
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5780..5797)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5951..6539)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6031..6050)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6710..7501,7502..7570))
                     /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     7333..7352
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7571..7675)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttacca tggtcccctc ggctggacag ctcgccctgt tcgctctggg
      961 tattgtgttg gctgcgtgcc aggccttgga gaacagcacg tccccgctgc gtacgatggt
     1021 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1081 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     1141 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1201 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     1261 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     1321 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     1381 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     1441 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     1501 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     1561 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     1621 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     1681 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtccggccg
     1741 gactcaggga gcagaattca tcgaaatcgt ggaaagatcc aactacatgg gtaacccatg
     1801 gactgagtac atggcaaagt acgacatcga ggaagtgcac ggaagtggta tcagggttga
     1861 tctgggtgaa gatgccgaag tcgctggtac tcagtacaga ctcccttctg gtaagtgccc
     1921 tgttttcgga aagggtatca tcatcgaaaa ctctaagact actttcctca agcctgttgc
     1981 tactggtaac caagatctta aggacggagg tttcgctttc ccacctacta accctctgat
     2041 ctctccaatg actttgaacg gtatgcgtga cttctacaag aacaacgaat acgtcaagaa
     2101 cttggatgaa ttgactttgt gtagtagaca cgctggaaac atgaaccctg ataacgacaa
     2161 gaacagtaac tacaagtacc ccgcggttta cgactacaac gataagaagt gtcacatcct
     2221 gtacatcgct gcccaagaaa acaacggacc aagatactgt aacaaggatc aaagtaagag
     2281 aaactctatg ttctgtttca gacctgcaaa ggacaagctg ttcgaaaact acgtgtactt
     2341 gtccaagaac gttgtcgata actgggaaga agtctgccca agaaagaacc tcgagaacgc
     2401 aaagttcggt ctgtgggtcg atggtaactg tgaagacatc cctcatgtga acgagttcag
     2461 tgctaacgat ttgttcgagt gtaacaagct ggtcttcgag ttgtctgcca gtgaccaacc
     2521 taagcagtac gaacagcatt tgactgacta cgaaaagatc aaggaaggat tcaagaacaa
     2581 gaacgccgat atgatcaagt ccgctttcct cccaaccggt gcattcaaag cagatagata
     2641 caagtctcac ggtaagggtt acaactgggg aaactacaac agagaaaccc aaaagtgtga
     2701 aatcttcaac gtcaagccta cctgcctcat caacgacaag tcctacattg cgactactgc
     2761 cctgtctcat ccaatcgaag tcgaacacaa cttcccctgc agtctctaca aggacgagat
     2821 caagaaggaa atcgagcgtg aaagtaagcg tatcaagttg aacgataacg acgacgaagg
     2881 taacaagaag atcatcgcac ctaggatctt catctccgat gacaaggatt ccctcaagtg
     2941 tccttgtgac cctgagatgg tgagtcagtc cacttgtaga ttcttcgttt gcaagtgcgt
     3001 cgaacgtaga gccgaagtca ctagtaacaa cgaagttgtc gtgaaggaag aatacaagga
     3061 tgaatacgct gatattccag agcataagcc tacgtacgat aaaatgatgc cgtattatgc
     3121 cggtgccggt gttctgttca tcattctggt gattctgggt gcaagccagg caaaatatca
     3181 gcgcctggag aaaatcaaca agaacaagat tgagaaaaac gtgaatgatt acaaggacga
     3241 tgacgataag taggcggccg cataatctag agggcccgtt taaacccgct gatcagcctc
     3301 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3361 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3421 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     3481 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     3541 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     3601 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     3661 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     3721 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     3781 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     3841 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     3901 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     3961 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     4021 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     4081 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4141 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     4201 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     4261 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
     4321 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
     4381 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     4441 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     4501 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     4561 accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg
     4621 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     4681 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     4741 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     4801 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     4861 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     4921 ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     4981 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     5041 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     5101 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     5161 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     5221 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     5281 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     5341 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     5401 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     5461 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     5521 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     5581 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     5641 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     5701 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     5761 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     5821 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5881 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5941 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     6001 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     6061 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     6121 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     6181 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6241 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     6301 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     6361 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     6421 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6481 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6541 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6601 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6661 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6721 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6781 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6841 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6901 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6961 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     7021 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     7081 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     7141 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     7201 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     7261 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     7321 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     7381 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     7441 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     7501 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     7561 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7621 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     7681 ttccccgaaa agtgccacct gacgtc
//
