LOCUS       sequence-139498-        7625 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Hairpin targeting GFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7625)
  AUTHORS   Jensen KJ, Moyer CB, Janes KA
  TITLE     Network Architecture Predisposes an Enzyme to Either Pharmacologic 
            or Genetic Targeting.
  JOURNAL   Cell Syst. 2016 Feb 24;2(2):112-121.
  PUBMED    26942229
REFERENCE   2  (bases 1 to 7625)
  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 
            Syst."; date: "2016-02-24"; volume: "2(2)"; pages: "112-121"
COMMENT     Sequence Label: shGFP neo
FEATURES             Location/Qualifiers
     source          1..7625
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     426..444
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             880..1674
                     /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(934..953)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1544..1563
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1696..1712)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1697..1713)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             1795..2028
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2100..2221
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2137..2156)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2191..2210
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2261..2396
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2416..2435)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2417..2435)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2445..2462)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2445..2461)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2454..2476)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2603..3058
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2690..2709)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2900..2921
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3084..3188
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3189..3257,3258..4049)
                     /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     complement(3407..3426)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4220..4808
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4709..4728
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4962..4979
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5096..5117
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5132..5137,5138..5155,5156..5162)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5170..5186
                     /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)."
     primer_bind     5175..5197
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5194..5210
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5194..5210
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5229..5249
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5231..5249
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5277..5503
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5504..5684
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5731..5856
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6349..6582
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6767..6811
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        7109..7357
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7109..7129
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7280..7299
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    7456..7573
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        join(7622..7625,1..507)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
ORIGIN
        1 ttggggttgc gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg
       61 cgtggttccg ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt
      121 cgcagcgtca cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct
      181 ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa
      241 gccgcacgtc tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc
      301 cgaccgcgat gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc
      361 cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc
      421 gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt
      481 gaccgaatca ccgacctctc tccccagggg gatccaccgg atcaagcttc acgctgccgc
      541 aagcactcag ggcgcaaggg ctgctaaagg aagcggaaca cgtagaaagc cagtccgcag
      601 aaacggtgct gaccccggat gaatgtcagc tactgggcta tctggacaag ggaaaacgca
      661 agcgcaaaga gaaagcaggt agcttgcagt gggcttacat ggcgatagct agactgggcg
      721 gttttatgga cagcaagcga accggaattg ccagctgggg cgccctctgg taaggttggg
      781 aagccctgca aagtaaactg gatggctttc ttgccgccaa ggatctgatg gcgcagggga
      841 tcaagatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
      901 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
      961 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     1021 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     1081 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     1141 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     1201 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     1261 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     1321 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     1381 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     1441 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     1501 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     1561 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     1621 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     1681 ctctggggtt cgcatcgata ccgtcgacct cgaggggggg cccggtacct ttaagaccaa
     1741 tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg ggactggaag
     1801 ggctaattca ctcccaacga agacaagatc tgctttttgc ttgtactggg tctctctggt
     1861 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
     1921 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
     1981 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt agtagttcat
     2041 gtcatcttat tattcagtat ttataacttg caaagaaatg aatatcagag agtgagagga
     2101 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     2161 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     2221 atcatgtctg gctctagcta tcccgcccct aactccgccc atcccgcccc taactccgcc
     2281 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     2341 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     2401 gacgtaccca attcgcccta tagtgagtcg tattacgcgc gctcactggc cgtcgtttta
     2461 caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc
     2521 cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg
     2581 cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     2641 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     2701 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     2761 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     2821 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     2881 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     2941 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     3001 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag
     3061 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     3121 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     3181 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     3241 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     3301 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     3361 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     3421 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     3481 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     3541 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     3601 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     3661 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     3721 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     3781 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     3841 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     3901 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     3961 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     4021 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     4081 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     4141 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     4201 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     4261 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     4321 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     4381 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     4441 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     4501 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     4561 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     4621 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     4681 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     4741 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     4801 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     4861 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     4921 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     4981 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     5041 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     5101 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     5161 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
     5221 ccaagcgcgc aattaaccct cactaaaggg aacaaaagct ggagctgcaa gcttaatgta
     5281 gtcttatgca atactcttgt agtcttgcaa catggtaacg atgagttagc aacatgcctt
     5341 acaaggagag aaaaagcacc gtgcatgccg attggtggaa gtaaggtggt acgatcgtgc
     5401 cttattagga aggcaacaga cgggtctgac atggattgga cgaaccactg aattgccgca
     5461 ttgcagagat attgtattta agtgcctagc tcgatacata aacgggtctc tctggttaga
     5521 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
     5581 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
     5641 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg
     5701 gacttgaaag cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa
     5761 gcgcgcacgg caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc
     5821 ggaggctaga aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga
     5881 tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat
     5941 atagtatggg caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca
     6001 tcagaaggct gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa
     6061 gaacttagat cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag
     6121 ataaaagaca ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc
     6181 accgcacagc aagcggccgc tgatcttcag acctggagga ggagatatga gggacaattg
     6241 gagaagtgaa ttatataaat ataaagtagt aaaaattgaa ccattaggag tagcacccac
     6301 caaggcaaag agaagagtgg tgcagagaga aaaaagagca gtgggaatag gagctttgtt
     6361 ccttgggttc ttgggagcag caggaagcac tatgggcgca gcgtcaatga cgctgacggt
     6421 acaggccaga caattattgt ctggtatagt gcagcagcag aacaatttgc tgagggctat
     6481 tgaggcgcaa cagcatctgt tgcaactcac agtctggggc atcaagcagc tccaggcaag
     6541 aatcctggct gtggaaagat acctaaagga tcaacagctc ctggggattt ggggttgctc
     6601 tggaaaactc atttgcacca ctgctgtgcc ttggaatgct agttggagta ataaatctct
     6661 ggaacagatt tggaatcaca cgacctggat ggagtgggac agagaaatta acaattacac
     6721 aagcttaata cactccttaa ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga
     6781 attattggaa ttagataaat gggcaagttt gtggaattgg tttaacataa caaattggct
     6841 gtggtatata aaattattca taatgatagt aggaggcttg gtaggtttaa gaatagtttt
     6901 tgctgtactt tctatagtga atagagttag gcagggatat tcaccattat cgtttcagac
     6961 ccacctccca accccgaggg gacccgacag gcccgaagga atagaagaag aaggtggaga
     7021 gagagacaga gacagatcca ttcgattagt gaacggatct cgacggtatc gatcacgaga
     7081 ctagcctcga gcggccgccc ccttcaccga gggcctattt cccatgattc cttcatattt
     7141 gcatatacga tacaaggctg ttagagagat aattggaatt aatttgactg taaacacaaa
     7201 gatattagta caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt
     7261 aaaattatgt tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt
     7321 cttggcttta tatatcttgt ggaaaggacg aaacaccggg caagctgacc ctgaagttca
     7381 tctgcagatg aacttcaggg tcagcttgct ttttgaattc tcgacctcga gacaaatggc
     7441 agtattcatc cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag
     7501 aatagtagac ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa
     7561 aattcaaaat tttcgggttt attacaggga cagcagagat ccactttggc cgcggctcga
     7621 ggggg
//
