LOCUS       sequence-211606-        3338 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express g-AAVS1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3338)
  AUTHORS   Ma S, Lv J, Sun J, Tang P, Li H, Zhou H, Zhang Z, Lin Y, Rong Z
  TITLE     iKA-CRISPR hESCs for inducible and multiplex orthogonal gene 
            knockout and activation.
  JOURNAL   FEBS Lett. 2018 Jun 5. doi: 10.1002/1873-3468.13127.
  PUBMED    29869798
REFERENCE   2  (bases 1 to 3338)
  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: "FEBS Lett. 
            2018 Jun 5. doi: 10.1002/1873-3468.13127."
COMMENT     Sequence Label: pU6-EF-gAAVS1
FEATURES             Location/Qualifiers
     source          1..3338
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     669..685
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     670..686
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             703..714
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        722..970
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     722..742
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     893..912
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        complement(1153..1171)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1192..1208)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1192..1208)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1205..1227)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1216..1232
                     /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)."
     promoter        complement(join(1240..1246,1247..1264,1265..1270))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1285..1306
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1423..1440)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1594..2182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1674..1693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2353..3144,3145..3213))
                     /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     2976..2995
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3214..3318)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tgggtaccgg
      661 gccccccctc gaggtcgacg gtatcgataa gcttgatatc gaattgaagg tcgggcagga
      721 agagggccta tttcccatga ttccttcata tttgcatata cgatacaagg ctgttagaga
      781 gataattaga attaatttga ctgtaaacac aaagatatta gtacaaaata cgtgacgtag
      841 aaagtaataa tttcttgggt agtttgcagt tttaaaatta tgttttaaaa tggactatca
      901 tatgcttacc gtaacttgaa agtatttcga tttcttggct ttatatatct tgtggaaagg
      961 acgaaacacc gtcaccaatc ctgtccctag gtttaagagc tatgctggaa acagcatagc
     1021 aagtttaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt
     1081 tttacgcgtt gacattgatt attgactagt tctagagcgg ccgccaccgc ggtggagctc
     1141 cagcttttgt tccctttagt gagggttaat ttcgagcttg gcgtaatcat ggtcatagct
     1201 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     1261 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     1321 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     1381 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     1441 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     1501 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     1561 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     1621 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     1681 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     1741 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     1801 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     1861 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     1921 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     1981 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     2041 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     2101 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2161 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2221 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     2281 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     2341 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     2401 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     2461 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     2521 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     2581 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     2641 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     2701 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     2761 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     2821 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     2881 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     2941 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     3001 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     3061 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     3121 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     3181 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     3241 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     3301 acaaataggg gttccgcgca catttccccg aaaagtgc
//
