LOCUS       Exported                7091 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pLKO.1 puro GFP siRNA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7091)
  AUTHORS   Orimo A, Gupta PB, Sgroi DC, Arenzana-Seisdedos F, Delaunay T, Naeem
            R, Carey VJ, Richardson AL, Weinberg RA
  TITLE     Stromal fibroblasts present in invasive human breast carcinomas 
            promote tumor growth and angiogenesis through elevated SDF-1/CXCL12 
            secretion.
  JOURNAL   Cell. 2005 May 6. 121(3):335-48.
  PUBMED    15882617
REFERENCE   2  (bases 1 to 7091)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7091
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        106..210
                     /gene="bla"
                     /label=AmpR promoter
     CDS             211..1071
                     /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(429..448)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1242..1830
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1731..1750
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1984..2001
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2118..2139
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        2154..2184
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2192..2208
                     /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     2197..2219
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2216..2232
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2216..2232
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2251..2271
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2253..2271
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2299..2525
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             2526..2706
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    2753..2878
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    3371..3604
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             3789..3833
                     /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        4131..4371
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     4131..4151
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     4302..4321
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    4483..4600
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        4649..5159
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     5078..5096
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             5181..5780
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5181..5200)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5677..5697
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5908..6141
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6213..6334
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6250..6269)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6304..6323
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6374..6509
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6436..6455
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(6529..6548)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6530..6548)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6558..6575)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6558..6574)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6567..6589)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6716..80
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6803..6822)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7013..7034
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
       61 aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt
      121 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
      181 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
      241 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
      301 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
      361 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
      421 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
      481 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
      541 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
      601 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
      661 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
      721 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
      781 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
      841 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
      901 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
      961 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     1021 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     1081 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     1141 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     1201 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     1261 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     1321 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     1381 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     1441 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     1501 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     1561 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     1621 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     1681 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     1741 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     1801 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     1861 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     1921 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     1981 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     2041 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     2101 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
     2161 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
     2221 aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag
     2281 ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc aacatggtaa
     2341 cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc cgattggtgg
     2401 aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg acatggattg
     2461 gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta gctcgataca
     2521 taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc taactaggga
     2581 acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc
     2641 tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct
     2701 ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg agctctctcg
     2761 acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc gactggtgag
     2821 tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg agagcgtcag
     2881 tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa
     2941 gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac gattcgcagt
     3001 taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac agctacaacc
     3061 atcccttcag acaggatcag aagaacttag atcattatat aatacagtag caaccctcta
     3121 ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca agatagagga
     3181 agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc agacctggag
     3241 gaggagatat gagggacaat tggagaagtg aattatataa atataaagta gtaaaaattg
     3301 aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga gaaaaaagag
     3361 cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc actatgggcg
     3421 cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata gtgcagcagc
     3481 agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc acagtctggg
     3541 gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag gatcaacagc
     3601 tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg ccttggaatg
     3661 ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg atggagtggg
     3721 acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa tcgcaaaacc
     3781 agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt ttgtggaatt
     3841 ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata gtaggaggct
     3901 tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt aggcagggat
     3961 attcaccatt atcgtttcag acccacctcc caaccccgag gggacccgac aggcccgaag
     4021 gaatagaaga agaaggtgga gagagagaca gagacagatc cattcgatta gtgaacggat
     4081 ctcgacggta tcgatcacga gactagcctc gagcggccgc ccccttcacc gagggcctat
     4141 ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag ataattggaa
     4201 ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat
     4261 ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg
     4321 taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacaccg
     4381 ggcccgcaag ctgaccctga agttcattca agagatgaac ttcagggtca gcttgctttt
     4441 tgaattctcg acctcgagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     4501 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     4561 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     4621 cagagatcca ctttggccgc ggctcgaggg ggttggggtt gcgccttttc caaggcagcc
     4681 ctgggtttgc gcagggacgc ggctgctctg ggcgtggttc cgggaaacgc agcggcgccg
     4741 accctgggtc tcgcacattc ttcacgtccg ttcgcagcgt cacccggatc ttcgccgcta
     4801 cccttgtggg ccccccggcg acgcttcctg ctccgcccct aagtcgggaa ggttccttgc
     4861 ggttcgcggc gtgccggacg tgacaaacgg aagccgcacg tctcactagt accctcgcag
     4921 acggacagcg ccagggagca atggcagcgc gccgaccgcg atgggctgtg gccaatagcg
     4981 gctgctcagc agggcgcgcc gagagcagcg gccgggaagg ggcggtgcgg gaggcggggt
     5041 gtggggcggt agtgtgggcc ctgttcctgc ccgcgcggtg ttccgcattc tgcaagcctc
     5101 cggagcgcac gtcggcagtc ggctccctcg ttgaccgaat caccgacctc tctccccagg
     5161 gggatccacc ggagcttacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
     5221 acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
     5281 gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
     5341 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
     5401 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
     5461 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
     5521 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
     5581 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
     5641 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
     5701 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
     5761 cccgcaagcc cggtgcctga cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg
     5821 cacgacccca tgcatcggta cctttaagac caatgactta caaggcagct gtagatctta
     5881 gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag
     5941 atctgctttt tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct
     6001 ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag
     6061 tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt
     6121 cagtgtggaa aatctctagc agtagtagtt catgtcatct tattattcag tatttataac
     6181 ttgcaaagaa atgaatatca gagagtgaga ggaacttgtt tattgcagct tataatggtt
     6241 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     6301 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggctctag ctatcccgcc
     6361 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     6421 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     6481 gaagtagtga ggaggctttt ttggaggcct agggacgtac ccaattcgcc ctatagtgag
     6541 tcgtattacg cgcgctcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc
     6601 gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa
     6661 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg
     6721 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     6781 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     6841 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     6901 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     6961 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     7021 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     7081 attttgccga t
//
