LOCUS       sequence-331139-        5666 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SFTPC reporter targeting repair template.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5666)
  AUTHORS   Sun D, Evans L, Perrone F, Sokleva V, Lim K, Rezakhani S, Lutolf M, 
            Zilbauer M, Rawlins EL
  TITLE     A functional genetic toolbox for human tissue-derived organoids.
  JOURNAL   eLife. 2021 Oct 6;10. pii: 67886. doi: 10.7554/eLife.67886.
  PUBMED    34612202
REFERENCE   2  (bases 1 to 5666)
  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: "eLife."; 
            date: "2021-10-6"; volume: "10. pii: 67886. doi"; pages: " 
            10.7554/eLife.67886"
COMMENT     Sequence Label: SFTPC-T2A-Rox-EF1a-Rox-Venus-NLS repair template
FEATURES             Location/Qualifiers
     source          1..5666
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             complement(1796..1816)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(join(1817..2527,2528..2530))
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1859..1880)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2208..2227
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2467..2488
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     protein_bind    2537..2568
                     /label=rox
                     /bound_moiety="Dre recombinase"
     LTR             2600..2868
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     promoter        complement(2881..3092)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     protein_bind    3125..3156
                     /label=rox
                     /bound_moiety="Dre recombinase"
     CDS             complement(3164..3217)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     complement(4041..4058)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4041..4057)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4050..4072)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4266..4285)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4385..4407
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4445..4463)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4531..4635
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4636..4704,4705..5496)
                     /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(4854..4873)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga cgacacgctg tcaggtcctc gcgccgcccc
     1021 tgccccgcgc ccccgcctcg ggtcggtctc tccgcccgcg tctccctatc tccccttctt
     1081 ggaccctcgc tctctcgcgt ctcgctctct gcgctctgtc tctctcgctg tcttggtgtc
     1141 tctccctcct ccggagacgc cgctggaggg cgccgaaagc ccgtggatgg ctccctgcga
     1201 gcccctctcc cccgccgagg tggacccagt gccccagcgc cctgggacag agggcgaatg
     1261 gcaaacccca ctttgctctc ccaaggccct tcccttcttg gccggccccc gccagggcag
     1321 gactgcgccc tgcaggaaga cagaagcaga ccctttgctt ttcaatcagg ctgctttatt
     1381 ctgttgtggc aggagttccc gggcccctgg gcgccacctc tgcccactcc catcctctcc
     1441 ccaagctccc atttctccag ggtttgtccc tgcggtgtgg gcagaagcag cttcttgccc
     1501 gtctgcaaaa gctgcaaaag accctttgcc cggggcgttt cctttcccgt tggggcttcc
     1561 actgaccctg tgaagcaaag gaatgttgag gatggacaga gagaattcag gggagtggga
     1621 agtaccggtc tgtgagcttc ccctcccccg agttgcctgg atgaccccgc ttcagtggac
     1681 gccccagcgc ctggtcagcg cgagcgctcc tccacagccc gggcaggtgg cccgagcgct
     1741 cctccgctgc tgatcagggg ccctgggatc acagctgctc agcggaggcg tcttatacct
     1801 ttctcttctt ttttggcttg tacagctcgt ccatgccgag agtgatcccg gcggcggtca
     1861 cgaactccag caggaccatg tgatcgcgct tctcgttggg gtctttgctc agggcggact
     1921 ggtagctcag gtagtggttg tcgggcagca gcacggggcc gtcgccgatg ggggtgttct
     1981 gctggtagtg gtcggcgagc tgcacgccgc cgtcctcgat gttgtggcgg atcttgaagt
     2041 tggccttgat gccgttcttc tgcttgtcgg cggtgatata gacgttgtgg ctgttgtagt
     2101 tgtactccag cttgtgcccc aggatgttgc cgtcctcctt gaagtcgatg cccttcagct
     2161 cgatgcggtt caccagggtg tcgccctcga acttcacctc ggcgcgggtc ttgtagttgc
     2221 cgtcgtcctt gaagaagatg gtgcgctcct ggacgtagcc ttcgggcatg gcggacttga
     2281 agaagtcgtg ctgcttcatg tggtcggggt agcgggcgaa gcactgcagg ccgtagccca
     2341 gggtggtcac gagggtgggc cagggcacgg gcagcttgcc ggtggtgcag atcagcttca
     2401 gggtcagctt gccgtaggtg gcatcgccct cgccctcgcc ggacacgctg aacttgtggc
     2461 cgtttacgtc gccgtccagc tcgaccagga tgggcaccac cccggtgaac agctcctcgc
     2521 ccttgctcac ggatcctaac tttaaataat tggcattatt taaagttatc atggtggcgt
     2581 aggcgccggt cacagcttgg atctgtaacg gcgcagaaca gaaaacgaaa caaagacgta
     2641 gagttgagca agcagggtca ggcaaagcgt ggagagccgg ctgagtctag gtaggctcca
     2701 agggagcgcc ggacaaaggc ccggtctcga cctgagcttt aaacttacct agacggcgga
     2761 cgcagttcag gaggcaccac aggcgggagg cggcagaacg cgactcaacc ggcgtggatg
     2821 gcggcctcag gtagggcggc gggcgcgtga aggagagatg cgagcccctc gaagcttcag
     2881 ctgtgttctg gcggcaaacc cgttgcgaaa aagaacgttc acggcgacta ctgcacttat
     2941 atacggttct cccccaccct cgggaaaaag gcggagccag tacacgacat cactttccca
     3001 gtttaccccg cgccaccttc tctaggcacc cgttcaattg ccgacccctc cccccaactt
     3061 ctcggggact gtgggcgatg tgcgctctgc ccactgacgg gcaccggagc gatcgcagat
     3121 cctttaactt taaataattg gcattattta aagttatacc ggtagggccg ggattctcct
     3181 ccacgtcacc gcatgttaga agacttcctc tgccctctcc ggatccgatg tagtacaacg
     3241 gcacctcgcc acacagggtg ctcacggcca tgcccaggaa ggccgggtcc cctccggagg
     3301 gtgctgagcc tgcatctcgc ccctctgcct ggcccagctt agacgtaggc actgcgggct
     3361 tggcctgcag agagcattcc atctggaatg taggaagtga gtttctatcc catgtgcagc
     3421 ccttattgtg ggacttcagg tatatcaggc aatctctctg agcctcagtt ttctcatcag
     3481 taaagagggg ctgtgctaga gtcagaagcc accaggtagt gagtggactc agctgagccc
     3541 agcagcttgg cagtggggtg ggaaggaaag gagtcgtgca ggggaatagg agaggggcag
     3601 tcagctggca gccaatgagg aacagtgctt tacaggtgac aaatagccac cattcggaca
     3661 ctcctcccag cagccctggg agggagacag cccactcttt tcacccacac acacacacct
     3721 ggaagttgtg gacttttcta gtgagagcct caagactggg gatgctctct ggagctatct
     3781 tcatgatgta gcagcaggtg ccaggggctg gcttgtaggc gatcagcagc taggcacaga
     3841 atgctggaga tcattcgggg gtccctgccc tgctctctcc ctgacccagg tccttagcca
     3901 cctccctcta ggttgggcac gggagtcata ctagaatctg gggctcaggc aaaaaaggtc
     3961 tggaggagag ggagagatgg atgtggatga agtggcccag ctgggtggct ctagcagagc
     4021 cttgtcattg gtcccagggt actggccgtc gttttacaac gtcgtgactg ggaaaaccct
     4081 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc
     4141 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc
     4201 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat atggtgcact
     4261 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     4321 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     4381 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     4441 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     4501 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     4561 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     4621 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     4681 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     4741 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     4801 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     4861 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     4921 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     4981 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     5041 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     5101 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     5161 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     5221 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     5281 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     5341 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     5401 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     5461 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     5521 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     5581 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     5641 cccgtagaaa agatcaaagg atcttc
//
