LOCUS       Exported                6031 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Mammalian expression plasmid for RBD of SARS-CoV-2 protein S (spike)
            with 8his tag.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3-SARS-CoV-2-S-RBD-8his
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6031)
  AUTHORS   Chan KK, Dorosky D, Sharma P, Abbasi SA, Dye JM, Kranz DM, Herbert 
            AS, Procko E
  TITLE     Engineering human ACE2 to optimize binding to the spike protein of 
            SARS coronavirus 2.
  JOURNAL   Science. 2020 Sep 4;369(6508):1261-1265. doi: 
            10.1126/science.abc0870. Epub 2020 Aug 4.
  PUBMED    32753553
REFERENCE   2  (bases 1 to 6031)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6031
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        136..515
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        516..719
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     670..690
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     764..783
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        764..782
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1456..1479
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(1523..1540)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1529..1753
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1799..2227
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1886..1905)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2096..2117
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2236..2256)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2241..2570
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2421..2556
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2483..2502
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2637..3431
                     /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(2691..2710)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3301..3320
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3605..3726
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3642..3661)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3696..3715
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3775..3791)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3775..3791)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3788..3810)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3799..3815
                     /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(3823..3853)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3868..3889
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4006..4023)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4177..4765)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4257..4276)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4936..5796)
                     /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     5559..5578
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5797..5901)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5976..5995)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
       61 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
      121 ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca attacggggt
      181 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
      241 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
      301 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
      361 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
      421 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
      481 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
      541 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
      601 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
      661 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc
      721 tctggctaac tagagaaccc actgcttact ggcttatcga aattaatacg actcactata
      781 gggagaccca agctggctag cgccaccatg aagaccatca tcgccctgag ctacatcttc
      841 tgcctggtgt tcgccactaa tctttgtccg ttcggtgagg tttttaacgc gacaaggttc
      901 gctagtgtat atgcttggaa ccgaaagaga atctccaatt gcgtagctga ttactccgtt
      961 ctctataaca gtgcgtcctt ttcaaccttt aagtgttacg gcgtttctcc aacgaagctg
     1021 aatgatctct gttttacgaa cgtgtatgct gactctttcg ttatacgggg ggacgaagtg
     1081 agacagatag caccaggtca gactgggaag atagcggatt acaactataa gttgcccgat
     1141 gattttacgg ggtgcgtaat cgcatggaac tcaaacaacc tcgactccaa agtaggtggt
     1201 aattataatt acttgtatcg cctgtttcga aagagcaatt tgaagccttt tgagcgggat
     1261 atttcaaccg aaatttacca agcaggcagt acgccatgta acggagtaga gggatttaat
     1321 tgctactttc ctcttcaatc ttatggcttt caaccaacaa acggagtggg gtatcaacct
     1381 tatagagtgg tagtattgtc ctttgagctc ctccacgccc cggctacagt ttgtgggccc
     1441 aaaaagggat ccggtcacca tcaccaccac catcatcatt gataactcga gtctagaggg
     1501 cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     1561 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     1621 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     1681 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     1741 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     1801 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     1861 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     1921 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     1981 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     2041 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     2101 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     2161 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     2221 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     2281 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     2341 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     2401 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     2461 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
     2521 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
     2581 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
     2641 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     2701 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     2761 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     2821 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     2881 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     2941 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     3001 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     3061 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     3121 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     3181 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     3241 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     3301 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     3361 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     3421 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     3481 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     3541 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     3601 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     3661 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     3721 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     3781 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     3841 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3901 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     3961 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     4021 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     4081 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     4141 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     4201 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     4261 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     4321 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     4381 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     4441 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4501 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4561 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     4621 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     4681 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     4741 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     4801 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     4861 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     4921 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     4981 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     5041 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     5101 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     5161 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     5221 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     5281 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     5341 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     5401 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     5461 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     5521 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     5581 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     5641 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     5701 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     5761 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     5821 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     5881 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     5941 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
     6001 ttaagccagt atctgctccc tgcttgtgtg t
//
