LOCUS       sequence-383802-        7084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bovine Creb5 shRNA targeting the Creb5 3′UTR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  AUTHORS   Zhang CH, Gao Y, Jadhav U, Hung HH, Holton KM, Grodzinsky AJ, 
            Shivdasani RA, Lassar AB
  TITLE     Creb5 establishes the competence for Prg4 expression in articular 
            cartilage.
  JOURNAL   Commun Biol. 2021 Mar 12;4(1):332. doi: 10.1038/s42003-021-01857-0.
  PUBMED    33712729
REFERENCE   2  (bases 1 to 7084)
  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: "Commun 
            Biol."; date: "2021-03-12"; volume: "4(1):332. doi"; pages: " 
            10.1038/s42003-021-01857-0"
COMMENT     Sequence Label: Creb5 shRNA
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     106..125
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     359..376
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    493..514
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(529..534,535..552,553..559)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    567..583
                     /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     572..594
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     591..607
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     591..607
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     626..646
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        628..646
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        674..900
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             901..1081
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1128..1253
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1746..1979
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2164..2208
                     /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        2506..2754
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2506..2526
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2677..2696
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2851..2968
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        3017..3527
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3446..3464
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3549..4148
                     /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(3549..3568)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4045..4065
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4276..4509
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4581..4702
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4618..4637)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4672..4691
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4742..4877
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4897..4916)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4898..4916)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4926..4943)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4926..4942)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4935..4957)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5084..5539
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5171..5190)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5381..5402
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5565..5669
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5670..5738,5739..6530)
                     /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(5888..5907)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(6701..7084,1..205)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
       61 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
      121 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
      181 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
      241 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
      301 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
      361 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
      421 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
      481 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
      541 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
      601 ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctgga
      661 gctgcaagct taatgtagtc ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg
      721 agttagcaac atgccttaca aggagagaaa aagcaccgtg catgccgatt ggtggaagta
      781 aggtggtacg atcgtgcctt attaggaagg caacagacgg gtctgacatg gattggacga
      841 accactgaat tgccgcattg cagagatatt gtatttaagt gcctagctcg atacataaac
      901 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
      961 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
     1021 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
     1081 agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca
     1141 ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc
     1201 caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta
     1261 agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa
     1321 aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc
     1381 ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc
     1441 ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg
     1501 tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc
     1561 aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc tggaggagga
     1621 gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca
     1681 ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg
     1741 ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg
     1801 tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac
     1861 aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc
     1921 aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg
     1981 gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt
     2041 tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga
     2101 gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa
     2161 gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt
     2221 aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta
     2281 ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca
     2341 ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata
     2401 gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa cggatctcga
     2461 cggtatcgat cacgagacta gcctcgagcg gccgccccct tcaccgaggg cctatttccc
     2521 atgattcctt catatttgca tatacgatac aaggctgtta gagagataat tggaattaat
     2581 ttgactgtaa acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt
     2641 gggtagtttg cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact
     2701 tgaaagtatt tcgatttctt ggctttatat atcttgtgga aaggacgaaa caccgggcct
     2761 tcaagaagag ctgttgcctc gaggcaacag ctcttcttga aggctttttg aattctcgac
     2821 ctcgagacaa atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta
     2881 cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca
     2941 aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca gagatccact
     3001 ttggccgcgg ctcgaggggg ttggggttgc gccttttcca aggcagccct gggtttgcgc
     3061 agggacgcgg ctgctctggg cgtggttccg ggaaacgcag cggcgccgac cctgggtctc
     3121 gcacattctt cacgtccgtt cgcagcgtca cccggatctt cgccgctacc cttgtgggcc
     3181 ccccggcgac gcttcctgct ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt
     3241 gccggacgtg acaaacggaa gccgcacgtc tcactagtac cctcgcagac ggacagcgcc
     3301 agggagcaat ggcagcgcgc cgaccgcgat gggctgtggc caatagcggc tgctcagcag
     3361 ggcgcgccga gagcagcggc cgggaagggg cggtgcggga ggcggggtgt ggggcggtag
     3421 tgtgggccct gttcctgccc gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt
     3481 cggcagtcgg ctccctcgtt gaccgaatca ccgacctctc tccccagggg gatccaccgg
     3541 agcttaccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca
     3601 gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg
     3661 atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg
     3721 ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca
     3781 cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt
     3841 tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc
     3901 ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg
     3961 gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg
     4021 ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg
     4081 tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg
     4141 gtgcctgacg cccgccccac gacccgcagc gcccgaccga aaggagcgca cgaccccatg
     4201 catcggtacc tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
     4261 aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat ctgctttttg
     4321 cttgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     4381 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     4441 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     4501 tctctagcag tagtagttca tgtcatctta ttattcagta tttataactt gcaaagaaat
     4561 gaatatcaga gagtgagagg aacttgttta ttgcagctta taatggttac aaataaagca
     4621 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     4681 ccaaactcat caatgtatct tatcatgtct ggctctagct atcccgcccc taactccgcc
     4741 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     4801 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     4861 aggctttttt ggaggcctag ggacgtaccc aattcgccct atagtgagtc gtattacgcg
     4921 cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
     4981 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
     5041 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
     5101 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     5161 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     5221 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     5281 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     5341 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     5401 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     5461 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     5521 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     5581 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     5641 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     5701 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     5761 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     5821 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     5881 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     5941 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     6001 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     6061 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     6121 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     6181 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     6241 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     6301 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     6361 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     6421 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     6481 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     6541 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     6601 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     6661 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     6721 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     6781 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     6841 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     6901 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     6961 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     7021 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     7081 taca
//
