LOCUS       Exported                9169 bp ds-DNA     circular SYN 17-NOV-2021
DEFINITION  Transient mammalian expression of codon optimized SARS-CoV-2 Spike 
            protein (original Wuhan Hu1 sequence + D614G mutation).
ACCESSION   .
VERSION     .
KEYWORDS    CoV2-Spike-D614G
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9169)
  AUTHORS   Syed AM, Taha TY, Tabata T, Chen IP, Ciling A, Khalid MM, Sreekumar 
            B, Chen PY, Hayashi JM, Soczek KM, Ott M, Doudna JA
  TITLE     Rapid assessment of SARS-CoV-2 evolved variants using virus-like 
            particles.
  JOURNAL   Science. 2021 Nov 4:eabl6184. doi: 10.1126/science.abl6184.
  PUBMED    34735219
REFERENCE   2  (bases 1 to 9169)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 17, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9169
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             complement(529..1389)
                     /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     1152..1171
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1390..1494)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1569..1588)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1760..2139
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2140..2343
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2294..2314
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2388..2407
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2388..2406
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6285..6302)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6291..6515
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6561..6989
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6648..6667)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6858..6879
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6998..7018)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7003..7332
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7183..7318
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7245..7264
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             7399..8193
                     /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(7453..7472)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     8063..8082
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    8367..8488
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8404..8423)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8458..8477
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(8537..8553)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8537..8553)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8550..8572)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8561..8577
                     /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(8585..8615)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8630..8651
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8768..8785)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8939..358)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9019..9038)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
       61 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
      121 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
      181 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
      241 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
      301 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
      361 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
      421 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
      481 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
      541 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
      601 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
      661 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
      721 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
      781 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
      841 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
      901 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
      961 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     1021 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     1081 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     1141 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     1201 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     1261 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     1321 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     1381 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     1441 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     1501 tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc
     1561 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt
     1621 gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg
     1681 accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta
     1741 cgggccagat atacgcgttg acattgatta ttgactagtt attaatagta atcaattacg
     1801 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
     1861 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
     1921 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
     1981 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
     2041 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
     2101 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
     2161 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
     2221 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
     2281 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
     2341 gctctctggc taactagaga acccactgct tactggctta tcgaaattaa tacgactcac
     2401 tatagggaga cccaagctgg ctagcatgtt tgtcttcctg gtcctgctgc ctctggtctc
     2461 gtctcagtgc gtgaacctga ctactagaac ccagctgcct cctgcctata ctaactcctt
     2521 cacccgcggc gtgtactacc cagacaaggt gttccgcagc tccgtgctgc actccaccca
     2581 ggatctgttc ctgcccttct tcagcaacgt gacctggttc cacgccatcc acgtgagcgg
     2641 caccaatggc accaagcggt tcgacaatcc cgtgctgcca ttcaacgatg gcgtgtactt
     2701 cgcctccacc gagaagagca acatcatccg cggctggatc ttcggcacca ccctggactc
     2761 caagacccag agcctgctga tcgtgaacaa tgccaccaac gtggtcatca aggtgtgcga
     2821 gttccagttc tgcaatgatc cattcctggg cgtgtactac cacaagaaca ataagtcctg
     2881 gatggagagc gagttccgcg tgtacagctc cgccaacaat tgcaccttcg agtacgtgtc
     2941 ccagcccttc ctgatggacc tggagggcaa gcagggcaat ttcaagaacc tgcgcgagtt
     3001 cgtgttcaag aatatcgatg gctacttcaa gatctactcc aagcacaccc ccatcaacct
     3061 ggtgcgcgac ctgccacagg gcttcagcgc cctggagcca ctggtggatc tgccaatcgg
     3121 catcaacatc accaggttcc agaccctgct ggccctgcac cgcagctacc tgaccccagg
     3181 cgacagctcc agcggatgga ccgctggagc tgctgcctac tacgtgggct acctgcagcc
     3241 ccgcaccttc ctgctgaagt acaacgagaa tggcaccatc accgacgccg tggattgcgc
     3301 cctggatcca ctgtccgaga caaagtgcac cctgaagagc ttcaccgtgg agaagggcat
     3361 ctaccagacc tccaatttcc gcgtgcagcc aaccgagagc atcgtgcgct tccccaatat
     3421 caccaacctg tgcccattcg gcgaggtgtt caacgctacc aggttcgcca gcgtgtacgc
     3481 ttggaatcgc aagcgcatct ccaactgcgt ggccgactac agcgtgctgt acaactccgc
     3541 cagcttctcc accttcaagt gctacggcgt gtcccccacc aagctgaatg atctgtgctt
     3601 caccaacgtg tacgccgata gcttcgtgat caggggcgac gaggtgcgcc agatcgctcc
     3661 aggacagacc ggcaagatcg ctgactacaa ttacaagctg cccgacgatt tcaccggctg
     3721 cgtgatcgcc tggaactcca acaatctgga tagcaaagtg ggcggcaact acaattacct
     3781 gtaccgcctg ttccgcaagt ccaatctgaa gccattcgag cgcgacatct ccaccgagat
     3841 ctaccaggct ggaagcaccc catgcaatgg agtggagggc ttcaactgct acttccccct
     3901 gcagagctac ggcttccagc caaccaacgg agtgggatac cagccataca gggtggtggt
     3961 gctgtccttc gagctgctgc acgctccagc taccgtgtgc ggaccaaaga agagcaccaa
     4021 tctggtgaag aacaagtgcg tgaacttcaa tttcaacggc ctgaccggaa ccggcgtgct
     4081 gaccgagtcc aacaagaagt tcctgccatt ccagcagttc ggaagggaca tcgctgatac
     4141 caccgacgcc gtgcgcgacc cacagaccct ggagatcctg gatatcaccc catgctcctt
     4201 cggcggcgtg agcgtgatca ccccaggaac caataccagc aaccaggtgg ccgtgctgta
     4261 ccagggcgtg aattgcaccg aggtgccagt ggctatccac gctgatcagc tgaccccaac
     4321 ctggcgcgtg tacagcaccg gatccaacgt gttccagacc cgcgccggat gcctgatcgg
     4381 agctgagcac gtgaacaatt cctacgagtg cgacatccca atcggagctg gaatctgcgc
     4441 cagctaccag acccagacca actccccaag gagggctcgc agcgtggcca gccagtccat
     4501 catcgcctac accatgtccc tgggcgccga gaatagcgtg gcctacagca acaattccat
     4561 cgccatccca accaacttca ccatctccgt gaccaccgag atcctgcccg tgtccatgac
     4621 caagaccagc gtggactgca ccatgtacat ctgcggcgat tccaccgagt gcagcaacct
     4681 gctgctgcag tacggcagct tctgcaccca gctgaatcgc gccctgaccg gaatcgctgt
     4741 ggagcaggat aagaacaccc aggaggtgtt cgcccaggtg aagcagatct acaagacccc
     4801 cccaatcaag gacttcggcg gcttcaattt cagccagatc ctgcccgatc caagcaagcc
     4861 ctccaagcgc agcttcatcg aggacctgct gttcaacaag gtgaccctgg ccgatgccgg
     4921 cttcatcaag cagtacggcg attgcctggg cgacatcgct gcccgcgacc tgatctgcgc
     4981 ccagaagttc aatggcctga ccgtgctgcc accactgctg accgatgaga tgatcgctca
     5041 gtacacctcc gccctgctgg ccggaaccat caccagcgga tggaccttcg gcgctggagc
     5101 cgccctgcag atccccttcg ccatgcagat ggcctaccgc ttcaacggca tcggcgtgac
     5161 ccagaatgtg ctgtacgaga accagaagct gatcgccaat cagttcaact ccgccatcgg
     5221 caagatccag gactccctgt ccagcaccgc cagcgccctg ggcaagctgc aggatgtggt
     5281 gaatcagaac gcccaggccc tgaataccct ggtgaagcag ctgtccagca acttcggcgc
     5341 catctccagc gtgctgaatg atatcctgag ccgcctggac aaggtggagg ctgaggtgca
     5401 gatcgatagg ctgatcaccg gccgcctgca gtccctgcag acctacgtga cccagcagct
     5461 gatcagggct gctgagatca gggccagcgc caatctggct gctaccaaga tgtccgagtg
     5521 cgtgctggga cagagcaaga gggtggactt ctgcggcaag ggctaccacc tgatgtcctt
     5581 cccacagagc gccccacacg gagtggtgtt cctgcacgtg acctacgtgc cagcccagga
     5641 gaagaacttc accaccgctc cagctatctg ccacgatggc aaggctcact tcccacgcga
     5701 gggcgtgttc gtgtccaacg gcacccactg gttcgtgacc cagcgcaatt tctacgagcc
     5761 ccagatcatc accaccgaca ataccttcgt gagcggcaac tgcgacgtgg tcatcggaat
     5821 cgtgaacaat accgtgtacg atcccctgca gccagagctg gactccttca aggaggagct
     5881 ggataagtac ttcaagaatc acaccagccc cgacgtggat ctgggcgaca tctccggcat
     5941 caatgccagc gtggtgaaca tccagaagga gatcgaccgc ctgaacgagg tggccaagaa
     6001 tctgaacgag tccctgatcg atctgcagga gctgggcaag tacgagcagt acatcaagtg
     6061 gccatggtac atctggctgg gcttcatcgc cggcctgatc gccatcgtga tggtgaccat
     6121 catgctgtgc tgcatgacct cctgctgcag ctgcctgaag ggctgctgct cctgcggcag
     6181 ctgctgcaag ttcgatgagg acgatagcga gcccgtgctg aagggcgtca aactgcacta
     6241 tacatagctc gagtctagag ggcccgttta aacccgctga tcagcctcga ctgtgccttc
     6301 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     6361 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     6421 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     6481 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
     6541 gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     6601 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     6661 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     6721 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     6781 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     6841 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     6901 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     6961 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt
     7021 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     7081 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     7141 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     7201 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     7261 gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     7321 ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca
     7381 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     7441 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     7501 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     7561 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
     7621 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     7681 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     7741 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     7801 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     7861 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     7921 aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     7981 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     8041 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     8101 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     8161 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     8221 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     8281 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     8341 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat
     8401 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     8461 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga
     8521 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     8581 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     8641 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     8701 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     8761 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     8821 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     8881 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     8941 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     9001 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     9061 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     9121 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggt
//
