LOCUS       62056_23305        7716 bp DNA     circular SYN 26-MAY-2020
DEFINITION  Cloning vector sh-AFP, complete sequence.
ACCESSION   MN996871
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7716)
  AUTHORS   Han X, Yang J, Zeng F, Weng J, Zhang Y, Peng Q, Shen L, Ding S, Liu 
            K, Gao Y.
  TITLE     Programmable Synthetic Protein Circuits for the Identification and 
            Suppression of Hepatocellular Carcinoma
  JOURNAL   Mol Ther Oncolytics 17, 70-82 (2020)
  PUBMED    32322664
REFERENCE   2  (bases 1 to 7716)
  AUTHORS   Liu K, Yang J, Ding S.
  TITLE     Direct Submission
  JOURNAL   Submitted (27-JAN-2020) Second Department of Hepatobiliary Surgery, 
            Zhujiang Hospital, No. 253, Industrial Avenue, Haizhu District, 
            Guangzhou, Guangdong 510245, China
REFERENCE   3  (bases 1 to 7716)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Ther 
            Oncolytics 17, 70-82 (2020)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (27-JAN-2020) Second Department of Hepatobiliary Surgery, Zhujiang 
            Hospital, No. 253, Industrial Avenue, Haizhu District, Guangzhou, 
            Guangdong 510245, China"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7716
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..792
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     misc_feature    814..1402
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     LTR             1474..1707
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1785..1919
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      1946..2081
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        complement(2102..2120)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2130..2146)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      2288..2743
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2769..2873
                     /label=AmpR promoter
     CDS             2874..3731
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      3905..4493
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    4781..4802
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        4817..4847
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4855..4871
                     /label=lac operator
                     /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)."
     primer_bind     4879..4895
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        4916..4934
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4962..5188
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     misc_feature    5416..5541
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    6034..6267
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6452..6496
                     /codon_start=1
                     /label=gp41 peptide
                     /note="antigenic peptide corresponding to amino acids 655
                     to 669 of the HIV envelope protein gp41 (Lutje Hulsik et 
                     al., 2013)"
                     /translation="KNEQELLELDKWASL"
     promoter        6674..6914
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_feature    7019..7136
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     promoter        7191..7695
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
ORIGIN
        1 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
       61 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
      121 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
      181 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
      241 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
      301 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
      361 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
      421 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
      481 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac
      541 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
      601 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
      661 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
      721 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
      781 gacgagttct tctgatgaac gcgttaagtc gacaatcaac ctctggatta caaaatttgt
      841 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
      901 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
      961 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     1021 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     1081 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     1141 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     1201 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
     1261 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     1321 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     1381 tccctttggg ccgcctcccc gcgtcgactt taagaccaat gacttacaag gcagctgtag
     1441 atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac tcccaacgaa
     1501 gacaagatct gctttttgct tgtactgggt ctctctggtt agaccagatc tgagcctggg
     1561 agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc
     1621 ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct
     1681 tttagtcagt gtggaaaatc tctagcagta cgtatagtag ttcatgtcat cttattattc
     1741 agtatttata acttgcaaag aaatgaatat cagagagtga gaggaacttg tttattgcag
     1801 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     1861 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggctct
     1921 agctatcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     1981 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     2041 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctagggacgt acccaattcg
     2101 ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg
     2161 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     2221 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     2281 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     2341 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2401 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2461 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2521 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2581 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2641 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2701 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     2761 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2821 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2881 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2941 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     3001 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     3061 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     3121 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     3181 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     3241 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     3301 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     3361 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     3421 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     3481 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3541 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3601 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3661 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3721 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3781 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3841 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3901 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3961 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4021 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4081 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4141 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4201 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4261 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4321 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4381 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4441 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4501 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4561 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4621 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     4681 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     4741 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     4801 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     4861 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag cgcgcaatta
     4921 accctcacta aagggaacaa aagctggagc tgcaagctta atgtagtctt atgcaatact
     4981 cttgtagtct tgcaacatgg taacgatgag ttagcaacat gccttacaag gagagaaaaa
     5041 gcaccgtgca tgccgattgg tggaagtaag gtggtacgat cgtgccttat taggaaggca
     5101 acagacgggt ctgacatgga ttggacgaac cactgaattg ccgcattgca gagatattgt
     5161 atttaagtgc ctagctcgat acataaacgg gtctctctgg ttagaccaga tctgagcctg
     5221 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
     5281 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     5341 cttttagtca gtgtggaaaa tctctagcag tggcgcccga acagggactt gaaagcgaaa
     5401 gggaaaccag aggagctctc tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga
     5461 ggcgaggggc ggcgactggt gagtacgcca aaaattttga ctagcggagg ctagaaggag
     5521 agagatgggt gcgagagcgt cagtattaag cgggggagaa ttagatcgcg atgggaaaaa
     5581 attcggttaa ggccaggggg aaagaaaaaa tataaattaa aacatatagt atgggcaagc
     5641 agggagctag aacgattcgc agttaatcct ggcctgttag aaacatcaga aggctgtaga
     5701 caaatactgg gacagctaca accatccctt cagacaggat cagaagaact tagatcatta
     5761 tataatacag tagcaaccct ctattgtgtg catcaaagga tagagataaa agacaccaag
     5821 gaagctttag acaagataga ggaagagcaa aacaaaagta agaccaccgc acagcaagcg
     5881 gccgctgatc ttcagacctg gaggaggaga tatgagggac aattggagaa gtgaattata
     5941 taaatataaa gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag
     6001 agtggtgcag agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg
     6061 agcagcagga agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt
     6121 attgtctggt atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca
     6181 tctgttgcaa ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga
     6241 aagataccta aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg
     6301 caccactgct gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa
     6361 tcacacgacc tggatggagt gggacagaga aattaacaat tacacaagct taatacactc
     6421 cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga
     6481 taaatgggca agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt
     6541 attcataatg atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat
     6601 agtgaataga gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc
     6661 gaggggaccc agagagggcc tatttcccat gattccttca tatttgcata tacgatacaa
     6721 ggctgttaga gagataatta gaattaattt gactgtaaac acaaagatat tagtacaaaa
     6781 tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa
     6841 aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat
     6901 cttgtggaaa ggacgaggta ccgggctgac ctggctacca tatttctcga gaaatatggt
     6961 agccaggtca gctttttgaa ttctagatct tgagacaaat ggcagtattc atccacaatt
     7021 ttaaaagaaa aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag
     7081 caacagacat acaaactaaa gaattacaaa aacaaattac aaaaattcaa aattttcggg
     7141 tttattacag ggacagcaga gatccacttt ggcgccggct cgagggggcc cgggttgcgc
     7201 cttttccaag gcagccctgg gtttgcgcag ggacgcggct gctctgggcg tggttccggg
     7261 aaacgcagcg gcgccgaccc tgggtctcgc acattcttca cgtccgttcg cagcgtcacc
     7321 cggatcttcg ccgctaccct tgtgggcccc ccggcgacgc ttcctgctcc gcccctaagt
     7381 cgggaaggtt ccttgcggtt cgcggcgtgc cggacgtgac aaacggaagc cgcacgtctc
     7441 actagtaccc tcgcagacgg acagcgccag ggagcaatgg cagcgcgccg accgcgatgg
     7501 gctgtggcca atagcggctg ctcagcaggg cgcgccgaga gcagcggccg ggaaggggcg
     7561 gtgcgggagg cggggtgtgg ggcggtagtg tgggccctgt tcctgcccgc gcggtgttcc
     7621 gcattctgca agcctccgga gcgcacgtcg gcagtcggct ccctcgttga ccgaatcacc
     7681 gacctctctc cccaggggga tccaccggag cttacc
//
