LOCUS       sequence-166131-        7237 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7237)
  AUTHORS   Godar S, Ince TA, Bell GW, Feldser D, Donaher JL, Bergh J, Liu A, 
            Miu K, Watnick RS, Reinhardt F, McAllister SS, Jacks T, Weinberg RA
  TITLE     Growth-inhibitory and tumor- suppressive functions of p53 depend on 
            its repression of CD44 expression.
  JOURNAL   Cell. 2008 Jul 11. 134(1):62-73.
  PUBMED    18614011
REFERENCE   2  (bases 1 to 7237)
  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. 2008 
            Jul 11. 134(1):62-73."
COMMENT     Sequence Label: shCD44-2 pRRL
FEATURES             Location/Qualifiers
     source          1..7237
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        25..129
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(130..198,199..990)
                     /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(348..367)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1161..1749
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1650..1669
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1903..1920
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2089..2110
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2125..2130,2131..2148,2149..2155)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2163..2179
                     /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     2168..2190
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2187..2203
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2187..2203
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2222..2242
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2224..2242
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2270..2496
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             2497..2677
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    2724..2849
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    3342..3575
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             3760..3804
                     /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        4102..4350
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     4102..4122
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     4273..4292
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    4448..4565
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        4614..5124
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     5043..5061
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     regulatory      5140..5149
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(5146..5148,5149..5151,5152..5865)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(5146..5148,5149..5151,5152..5862)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5191..5212)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5452..5471)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5799..5820
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5973..6206
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6278..6399
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6315..6334)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6369..6388
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6439..6574
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6594..6613)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6595..6613)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6623..6640)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6623..6639)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6632..6654)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6781..7209
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6868..6887)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7078..7099
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
       61 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
      121 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
      181 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
      241 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
      301 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
      361 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
      421 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
      481 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
      541 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
      601 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
      661 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
      721 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
      781 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
      841 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
      901 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
      961 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     1021 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     1081 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     1141 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     1201 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     1261 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     1321 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     1381 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     1441 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     1501 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     1561 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     1621 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     1681 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     1741 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     1801 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     1861 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     1921 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     1981 tgagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     2041 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
     2101 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
     2161 aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagcg
     2221 cgcaattaac cctcactaaa gggaacaaaa gctggagctg caagcttaat gtagtcttat
     2281 gcaatactct tgtagtcttg caacatggta acgatgagtt agcaacatgc cttacaagga
     2341 gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg tgccttatta
     2401 ggaaggcaac agacgggtct gacatggatt ggacgaacca ctgaattgcc gcattgcaga
     2461 gatattgtat ttaagtgcct agctcgatac ataaacgggt ctctctggtt agaccagatc
     2521 tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg
     2581 ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc
     2641 ctcagaccct tttagtcagt gtggaaaatc tctagcagtg gcgcccgaac agggacttga
     2701 aagcgaaagg gaaaccagag gagctctctc gacgcaggac tcggcttgct gaagcgcgca
     2761 cggcaagagg cgaggggcgg cgactggtga gtacgccaaa aattttgact agcggaggct
     2821 agaaggagag agatgggtgc gagagcgtca gtattaagcg ggggagaatt agatcgcgat
     2881 gggaaaaaat tcggttaagg ccagggggaa agaaaaaata taaattaaaa catatagtat
     2941 gggcaagcag ggagctagaa cgattcgcag ttaatcctgg cctgttagaa acatcagaag
     3001 gctgtagaca aatactggga cagctacaac catcccttca gacaggatca gaagaactta
     3061 gatcattata taatacagta gcaaccctct attgtgtgca tcaaaggata gagataaaag
     3121 acaccaagga agctttagac aagatagagg aagagcaaaa caaaagtaag accaccgcac
     3181 agcaagcggc cgctgatctt cagacctgga ggaggagata tgagggacaa ttggagaagt
     3241 gaattatata aatataaagt agtaaaaatt gaaccattag gagtagcacc caccaaggca
     3301 aagagaagag tggtgcagag agaaaaaaga gcagtgggaa taggagcttt gttccttggg
     3361 ttcttgggag cagcaggaag cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc
     3421 agacaattat tgtctggtat agtgcagcag cagaacaatt tgctgagggc tattgaggcg
     3481 caacagcatc tgttgcaact cacagtctgg ggcatcaagc agctccaggc aagaatcctg
     3541 gctgtggaaa gatacctaaa ggatcaacag ctcctgggga tttggggttg ctctggaaaa
     3601 ctcatttgca ccactgctgt gccttggaat gctagttgga gtaataaatc tctggaacag
     3661 atttggaatc acacgacctg gatggagtgg gacagagaaa ttaacaatta cacaagctta
     3721 atacactcct taattgaaga atcgcaaaac cagcaagaaa agaatgaaca agaattattg
     3781 gaattagata aatgggcaag tttgtggaat tggtttaaca taacaaattg gctgtggtat
     3841 ataaaattat tcataatgat agtaggaggc ttggtaggtt taagaatagt ttttgctgta
     3901 ctttctatag tgaatagagt taggcaggga tattcaccat tatcgtttca gacccacctc
     3961 ccaaccccga ggggacccga caggcccgaa ggaatagaag aagaaggtgg agagagagac
     4021 agagacagat ccattcgatt agtgaacgga tctcgacggt atcgatcacg agactagcct
     4081 cgagcggccg cccccttcac cgagggccta tttcccatga ttccttcata tttgcatata
     4141 cgatacaagg ctgttagaga gataattgga attaatttga ctgtaaacac aaagatatta
     4201 gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta
     4261 tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct
     4321 ttatatatct tgtggaaagg acgaaacacc ggtatgcaat gtgctactga ttgtctcgag
     4381 acaatcagta gcacattgca ttttttgaat tctcgacctc gagacaaatg gcagtattca
     4441 tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag
     4501 acataatagc aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa
     4561 attttcgggt ttattacagg gacagcagag atccactttg gccgcggctc gagggggttg
     4621 gggttgcgcc ttttccaagg cagccctggg tttgcgcagg gacgcggctg ctctgggcgt
     4681 ggttccggga aacgcagcgg cgccgaccct gggtctcgca cattcttcac gtccgttcgc
     4741 agcgtcaccc ggatcttcgc cgctaccctt gtgggccccc cggcgacgct tcctgctccg
     4801 cccctaagtc gggaaggttc cttgcggttc gcggcgtgcc ggacgtgaca aacggaagcc
     4861 gcacgtctca ctagtaccct cgcagacgga cagcgccagg gagcaatggc agcgcgccga
     4921 ccgcgatggg ctgtggccaa tagcggctgc tcagcagggc gcgccgagag cagcggccgg
     4981 gaaggggcgg tgcgggaggc ggggtgtggg gcggtagtgt gggccctgtt cctgcccgcg
     5041 cggtgttccg cattctgcaa gcctccggag cgcacgtcgg cagtcggctc cctcgttgac
     5101 cgaatcaccg acctctctcc ccagggggat ccaccggtcg ccaccatggt gagcaagggc
     5161 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     5221 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     5281 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     5341 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
     5401 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     5461 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     5521 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     5581 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     5641 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     5701 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag
     5761 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     5821 accgccgccg ggatcactct cggcatggac gagctgtaca agtaaagcgg ccgcgtcgac
     5881 gggcccgcgg aattcgagct cggtaccttt aagaccaatg acttacaagg cagctgtaga
     5941 tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag
     6001 acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga
     6061 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     6121 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     6181 ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt
     6241 ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa
     6301 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     6361 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc
     6421 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     6481 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     6541 ttccagaagt agtgaggagg cttttttgga ggcctaggga cgtacccaat tcgccctata
     6601 gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     6661 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     6721 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg
     6781 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     6841 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     6901 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     6961 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     7021 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     7081 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     7141 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     7201 acgcgaattt taacaaaata ttaacgctta caattta
//
