LOCUS       sequence-319220-        7753 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting vector to TH gene locus for induction of P2A-fused iCre, 
            with PGK-puroR drug resistant cassette flanked by FRT sequence..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7753)
  AUTHORS   Xiong M, Tao Y, Gao Q, Feng B, Yan W, Zhou Y, Kotsonis TA, Yuan T, 
            You Z, Wu Z, Xi J, Haberman A, Graham J, Block J, Zhou W, Chen Y, 
            Zhang SC
  TITLE     Human Stem Cell-Derived Neurons Repair Circuits and Restore Neural 
            Function.
  JOURNAL   Cell Stem Cell. 2020 Sep 16. pii: S1934-5909(20)30410-0. doi: 
            10.1016/j.stem.2020.08.014.
  PUBMED    32966778
REFERENCE   2  (bases 1 to 7753)
  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 Stem 
            Cell. 2020 Sep 16. pii: S1934-5909(20)30410-0. doi: 
            10.1016/j.stem.2020.08.014."
COMMENT     Sequence Label: Th-iCre
FEATURES             Location/Qualifiers
     source          1..7753
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..57
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             64..84
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    1123..1156
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     promoter        1217..1704
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1252..1275)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1637..1656
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             1744..2343
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APHNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1744..1763)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     protein_bind    2900..2933
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(3941..3960)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3942..3960)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3967..3984)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3967..3983)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3976..3998)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(4124..4579)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4261..4282)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4473..4492
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4606..4710
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4711..4779,4780..5571)
                     /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(4929..4948)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5742..6330
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6231..6250
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6484..6501
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6618..6639
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6654..6659,6660..6677,6678..6684)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6692..6708
                     /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     6697..6719
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6716..6732
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6716..6732
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        6753..6771
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 gcaacaaact tctctctgct gaaacaagcc ggagatgtcg aagagaatcc tggaccgatg
       61 gtgcccaaga agaagaggaa agtctccaac ctgctgactg tgcaccaaaa cctgcctgcc
      121 ctccctgtgg atgccacctc tgatgaagtc aggaagaacc tgatggacat gttcagggac
      181 aggcaggcct tctctgaaca cacctggaag atgctcctgt ctgtgtgcag atcctgggct
      241 gcctggtgca agctgaacaa caggaaatgg ttccctgctg aacctgagga tgtgagggac
      301 tacctcctgt acctgcaagc cagaggcctg gctgtgaaga ccatccaaca gcacctgggc
      361 cagctcaaca tgctgcacag gagatctggc ctgcctcgcc cttctgactc caatgctgtg
      421 tccctggtga tgaggagaat cagaaaggag aatgtggatg ctggggagag agccaagcag
      481 gccctggcct ttgaacgcac tgactttgac caagtcagat ccctgatgga gaactctgac
      541 agatgccagg acatcaggaa cctggccttc ctgggcattg cctacaacac cctgctgcgc
      601 attgccgaaa ttgccagaat cagagtgaag gacatctccc gcaccgatgg tgggagaatg
      661 ctgatccaca ttggcaggac caagaccctg gtgtccacag ctggtgtgga gaaggccctg
      721 tccctggggg ttaccaagct ggtggagaga tggatctctg tgtctggtgt ggctgatgac
      781 cccaacaact acctgttctg ccgggtcaga aagaatggtg tggctgcccc ttctgccacc
      841 tcccaactgt ccacccgggc cctggaaggg atctttgagg ccacccaccg cctgatctat
      901 ggtgccaagg atgactctgg gcagagatac ctggcctggt ctggccactc tgccagagtg
      961 ggtgctgcca gggacatggc cagggctggt gtgtccatcc ctgaaatcat gcaggctggt
     1021 ggctggacca atgtgaacat tgtgatgaac tacatcagaa acctggactc tgagactggg
     1081 gccatggtga ggctgctcga ggatggggac tgataggaat tcgaagttcc tattctctag
     1141 aaagtatagg aacttctagg tctgaagagg agtttacgtc cagccaagct taggatctcg
     1201 acctcgaaat tctaccgggt aggggaggcg cttttcccaa ggcagtctgg agcatgcgct
     1261 ttagcagccc cgctgggcac ttggcgctac acaagtggcc tctggcctcg cacacattcc
     1321 acatccaccg gtaggcgcca accgactccg ttctttggtg gccccttcgc gccaccttct
     1381 actcctcccc tagtcaggaa gttccccccc gccccgcagc tcgcgtcgtg caggacgtga
     1441 caaatggaag tagcacgtct cactagtctc gtgcagatgg acagcaccgc tgagcaatgg
     1501 aagcgggtag gcctttgggg cagcggccaa tagcagcttt gctccttcgc tttctgggct
     1561 cagaggctgg gaaggggtgg gtccgggggc gggctcaggg gcggggcggg cgcccgaagg
     1621 tcctccggag gcccggcatt ctgcacgctt caaaagcgca cgtctgccgc gctgttctcc
     1681 tcttcctcat ctccgggcct ttcgacctgc atccatctag atctcgagca gctgaagctt
     1741 accatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc
     1801 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg
     1861 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
     1921 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
     1981 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
     2041 ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag
     2101 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
     2161 ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc
     2221 ctggagacct ccgcgcccca caacctcccc ttctacgagc ggctcggctt caccgtcacc
     2281 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
     2341 tgacgcccgc cccacgaccc gcagcgcccg accgaaagga gcgcacgacc ccatgcatcg
     2401 atgatatcag atccccggga tgcagaaatt gatgatctat taaacaataa agatgtccac
     2461 taaaatggaa gtttttcctg tcatactttg ttaagaaggg tgagaacaga gtacctacat
     2521 tttgaatgga aggattggag ctacgggggt gggggtgggg tgggattaga taaatgcctg
     2581 ctctttactg aaggctcttt actattgctt tatgataatg tttcatagtt ggatatcata
     2641 atttaaacaa gcaaaaccaa attaagggcc agctcattcc tcccactcat gatctataga
     2701 tctatagatc tctcgtggga tcattgtttt tctcttgatt cccactttgt ggttctaagt
     2761 actgtggttt ccaaatgtgt cagtttcata gcctgaagaa cgagatcagc agcctctgtt
     2821 ccacatacac ttcattctca gtattgtttt gccaagttct aattccatca gaagctggtc
     2881 gagatccgga acccttaatg aagttcctat tctctagaaa gtataggaac ttcggatctt
     2941 aggtgcacgg cgtccctgaa ctcccttccc aacctcgttt ggtcctgcac tgtcccggaa
     3001 tccaggccct ggtgaggggc tgggtcccgg gtgcccccca tgccctccct gctgccaggc
     3061 tcccactgcc cctgcacctg cttctcagcg caacagctgt gtgtgcccgt ggtgaggttg
     3121 tgctgcctgt ggtgaggtcc tgtcctggct cccagggtcc tgggggctgc tgcactgccc
     3181 tccgcccttc cctgacactg tctgctgccc caatcaccgt cacaataaaa gaaactgtgg
     3241 tctctacacc tgcctggccc cacatctgtg ccacagagac agaccctggg atcctcagac
     3301 tcccacaccc ccaccccagc ctcactcaga ggtttcgccc tggcctcctt cctcctctgg
     3361 gagatggctg gccgccctgg ccaggcagct ggcccctccg ggcctggttt ccccgctcac
     3421 cctgaggccc cgcccagctc tgagccccaa gcagctccag aggctcgggc accctggccg
     3481 agctgcccca tctccgtggg gtgccctccc aaggtgggga gccacgtgac agtgggaggg
     3541 cctctctcag gcctggaagg gagcaggggt cacaaactgt gctggctggg ggtggtctca
     3601 gaggtgggcc tgcaggccta accctccctg ctgacagggc tcccagccct tgagagaaac
     3661 agggatggag gaacagctgc cctgatgccc tcacccaccc ggagcaggcc ctgcgaacca
     3721 aggggaacct cagtgtggcc cccagcatgt gtgctgatgg ggagggtctg gctgagctgg
     3781 tgcccaggca gatggtctgg gcctgtctcc ccagcgaggc aggatggggg ctggatttca
     3841 gactctgtaa gatgcccctg gcttactcga ggggcctgga cattgccctc cagagagagc
     3901 acccaacacg cggccgccac cgcggtggag ctccaattcg ccctatagtg agtcgtatta
     3961 caattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     4021 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     4081 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggaaattgt aagcgttaat
     4141 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     4201 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     4261 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     4321 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     4381 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     4441 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     4501 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     4561 gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     4621 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     4681 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     4741 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     4801 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     4861 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     4921 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     4981 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     5041 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     5101 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     5161 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     5221 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     5281 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     5341 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     5401 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     5461 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     5521 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     5581 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     5641 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     5701 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     5761 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     5821 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     5881 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     5941 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     6001 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     6061 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     6121 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     6181 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     6241 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     6301 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     6361 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     6421 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     6481 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     6541 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     6601 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
     6661 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
     6721 aacagctatg accatgatta cgccaagctc gaaattaacc ctcactaaag ggaacaaaag
     6781 ctggtaccgg ccagccaggc cttccttctg gccttgagca gggctggacc tgtgagccca
     6841 ggtcacagat gagaaaaccg acccctggct gcagcagccc ccacacagca gggacaccat
     6901 ccgtgagaag gaccccagcg tctggggagg ggcagaccta caggactggg ggctgctggg
     6961 tggccgggtc aaggccagtc ttggaggtgc tgacagagcc tgagctttgt gaggacgtcc
     7021 tgtggaacct gtcccggccc cctgccctgg gatggggaga agtcaggggg atagacagag
     7081 tcaaggtggg ggacagggcg ggagtggggt ccccagggct gggggccttt ggtgcagtga
     7141 ccagagtgtc aggagagggg agcaaagccc tctagcctca tcctcataaa aggtctcatc
     7201 attttccctc cagcctctta tgcactgggg aaactgaggc caggggctat gtgtccagcg
     7261 gacaggggtg ctgaattcca cccacaggct tagggatatg gtcaaggaaa gcttcctgga
     7321 ggaggcccag tggaggttca gggagggatg gggtgcccgg cagtctctag tggaaaaggc
     7381 gcctagccta tctcccccat gaaccccctc acccagccct ggaagaggcc tcagtgtccc
     7441 gcctgtgacc agttggctca gaaaagccct gggagctctg agccactgtg aaggtggaaa
     7501 cgcggcccct ggcctcccct ctcctggagg ctgcagactc tgcccgccag ttgacgaggg
     7561 ctctgccgct ctcctcccca ggagctatgc ctcacgcatc cagcgcccct tctccgtgaa
     7621 gttcgacccg tacacgctgg ccatcgacgt gctggacagc ccccaggccg tgcggcgctc
     7681 cctggagggt gtccaggatg agctggacac ccttgcccat gcgctgagtg ccattggcgt
     7741 cgacggatcc ggc
//
