LOCUS       sequence-273354-        8531 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express HIV Env which is isolated a from IRB approved HIV infected 
            patient samples from Singapore and cloned into expression vector 
            (pcDNA 3.1).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8531)
  AUTHORS   Joshi A, Cox EK, Sedano MJ, Punke EB, Lee RT, Maurer-Stroh S, Kaur 
            P, Ng OT, Garg H
  TITLE     HIV-1 subtype CRF01_AE and B differ in utilization of low levels of 
            CCR5, Maraviroc susceptibility and potential N-glycosylation sites.
  JOURNAL   Virology. 2017 Dec;512:222-233. doi: 10.1016/j.virol.2017.09.026. 
            Epub 2017 Oct 9.
  PUBMED    29020646
REFERENCE   2  (bases 1 to 8531)
  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: "Virology.";
            date: "2017-12"; volume: "512:222-233. doi"; pages: " 
            10.1016/j.virol.2017.09.026. Epub 2017 Oct 9"
COMMENT     Sequence Label: pFREE019Env
FEATURES             Location/Qualifiers
     source          1..8531
                     /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             4029..4070
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             4080..4097
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4120..4137)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4126..4350
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4396..4824
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4483..4502)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4693..4714
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4833..4853)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4838..5167
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5018..5153
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5080..5099
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5234..6028
                     /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(5288..5307)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5898..5917
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6204..6325
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6241..6260)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6295..6314
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6374..6390)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6374..6390)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6387..6409)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6398..6414
                     /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(6422..6428,6429..6446,6447..6452))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6467..6488
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6605..6622)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6776..7364)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6856..6875)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7535..8326,8327..8395))
                     /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     8158..8177
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8396..8500)
                     /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 gctggctagt
      901 taagcttggt accgagctcg gatccagtac ccttcaccga tcaagcttta ggcatctcct
      961 atggcaggaa gaagcggaag caccgacgag gaactcctca gagcagtaag gatcatcaaa
     1021 ctcctctacc aaagcagtaa gtaatatatg taatgtcacc cttggaaatt agtgcaataa
     1081 caggactagt aatagcgcta atcttagcaa tagtagtgtg gactatagta gctatagaag
     1141 ttaggaaaat actaaggcaa agaaaaatag acaggttaat taagagaata agagaaagag
     1201 cagaagacag tggaaatgag agtgaagggg acacagatga gttggccaaa cttgtggaaa
     1261 tgggggactt tgatccttgg gttggtgata atttgtagtg cctcagacaa cttgtgggtt
     1321 acagtttatt atggggtccc tgtatggaaa gatgcagata ccaccctatt ttgtgcatca
     1381 gatgccaaag cacaagagac agaagcgcac aatgtctggg ccacacatgc ctgtgtaccc
     1441 acagacccca acccacaaga aatatacatg gaaaatgtaa cagaaaattt taacatgtgg
     1501 aaaaataaca tggtagagca gatgcaggag gatgtaatca gtttatggga tcaaagtcta
     1561 cagccatgtg taaagctaac tcctctctgc gtaactttaa attgtactga tgctaaagtg
     1621 atccttgaca caaacacaac ccaagcccca agcacaaaca caaccaccac aaacacaacc
     1681 actgtcccta gagtaaacac aacagacgaa gtaaaaaact gttcttttaa tacaaccaca
     1741 gtaataagag ataagaagca gaaggtccat gcactttttt ataggcttga tatagtacca
     1801 atagataaca gtggtcaaat ggataacagt agtagtaggg agtataggtt aataaattgt
     1861 aatacttcag ttattaagca ggcttgtcca aaaatatcct ttgatccaat tcctatacat
     1921 tattgtactc ctgctggtta tgtgatttta aagtgtaatg ataagaagtt caatgggaca
     1981 gggccatgta gaaatgtcag ctcagtacaa tgcacacatg gaattaagcc agtggtatca
     2041 actcaattgc tgttaaatgg cagtctagct gaagaagata taataatcag atctgaaaat
     2101 ctcacaaaca atgcaaaaac cataatagtg caccttaata aatctgtaga aatcaattgt
     2161 accagaccct ccaacaatac aagaaaaggt acaactatag gaccaggaca agtattctat
     2221 agaacgggag aaataatagg agatataaga caagcacatt gtgagattaa tggaacaaaa
     2281 tggaaggaag tgttaacaca ggtagctaaa gaattacaaa agcactttaa caataagcca
     2341 ataaaatttc aaccaccctc aggaggagat ctagaaatta caactcatca tttcaattgt
     2401 agaggggaat ttttctattg caatacatca aacctgttta atagtactta taataacact
     2461 gagttaactg taggtaataa tactgataat gatactatca cgcttccatg caggataaag
     2521 caaattataa acatgtggca gggagcagga caagcaatgt atgctcctcc catcagtgga
     2581 aaaattaatt gtatatcaaa tattacagga atattattga caagagatgg tggtaataat
     2641 gcaacaaatg aagagatctt cagacctgaa ggaggaaata taaaggacaa ttggagaagt
     2701 gaattatata aatataaagt agtacaaatt gaaccactag gaatagcacc caccagggca
     2761 aagagaagag tggtggagag agaaaaaaga gcagtgggaa taggagctat gatctttggg
     2821 ttcttaggag cagccggaag cactatgggc gcagcgtcaa taacgctgac ggtacaggcc
     2881 agacaattat tgtctggtat agtgcaacag caaaacaatt tgctgagggc tatagaggcg
     2941 cagcagcata tgttgcaact cacagtctgg ggcattaaac agctccaggc aagagtcctg
     3001 gctgtggaaa gatacctaaa ggatcaaaag ttcctaggac tttggggctg ctctggaaaa
     3061 atcatctgca ccactgctgt gccctggaac tccacttgga gtaataaatc ttttgatgag
     3121 atttggaaca acatgacatg ggcagaatgg gaaagagaaa ttagcaatta cacaaaccaa
     3181 atatatgaca tacttacaga atcacagaac cagcaggaca ggaatgaaaa ggatttgtta
     3241 gaattagata aatggacaag tctgtggagt tggtttgaca taacaaattg gctgtggtat
     3301 ataaaaatat ttataatgat agtaggaggt ttaataggtt taagaataat ttttgctgtg
     3361 ctttctatag taaatagagt taggcagggt tactcacctt tgtctttcca gatccctacc
     3421 catcagcaga gggaaccaga caggcccgaa agaaccgaag aaggaggtgg cgagcaaggc
     3481 agagacagat ccgtgagatt agtgagcggg ttcttagcgc ttgcgtggga cgacctacgg
     3541 agcctgtgcc tcttcagcta ccacctcttg agaaacttca tctcgattgc agcgaggact
     3601 gtggaacttc tgggacgcag cagcctcaag ggactgagac gggggtggga aggtcttaaa
     3661 tatctaggga atcttctgtt atattggggc caggaactaa aaattagtgc tatctctttg
     3721 cttgatgcca cagcaataac aatagcgggg tggacagata ggattataga actagcacaa
     3781 ggagttggga gagccattct acatatacct aggagaatca gacaggggtt ggaaagggct
     3841 ttgctataac atgggaagca agtggtcaaa aagttggcct caggtcagag atagaataaa
     3901 gcaaactcct ccagcagcag aaggagtagg agcagtatct caagacggat ccagctaagg
     3961 gtcaagacaa ttctgcagat atccagcaca gtggcggccg ctcgagtcta gagggcccgc
     4021 ggttcgaagg taagcctatc cctaaccctc tcctcggtct cgattctacg cgtaccggtc
     4081 atcatcacca tcaccattga gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     4141 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     4201 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     4261 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     4321 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     4381 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     4441 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     4501 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     4561 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     4621 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     4681 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     4741 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     4801 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     4861 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4921 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4981 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     5041 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     5101 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     5161 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg
     5221 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     5281 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     5341 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     5401 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     5461 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     5521 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     5581 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     5641 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     5701 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     5761 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     5821 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     5881 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     5941 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     6001 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgcgaa atgaccgacc
     6061 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     6121 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     6181 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     6241 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     6301 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     6361 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     6421 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     6481 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     6541 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     6601 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6661 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6721 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6781 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6841 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     6901 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     6961 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     7021 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     7081 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     7141 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     7201 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     7261 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     7321 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     7381 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     7441 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     7501 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     7561 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     7621 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     7681 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     7741 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     7801 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     7861 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     7921 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     7981 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     8041 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     8101 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     8161 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     8221 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     8281 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     8341 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     8401 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     8461 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     8521 cacctgacgt c
//
