LOCUS       sequence-390930-        7658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LV Expression of CD3D cDNA with mutation at c.274+5G>A used as a 
            disease editing target for CD3d SCID mutation identified in Ecuador 
            .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7658)
  TITLE     Kohn lab plasmids
REFERENCE   2  (bases 1 to 7658)
  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"
COMMENT     Sequence Label: pCCL-c-MNDU3-CD3D-274+5G>A
FEATURES             Location/Qualifiers
     source          1..7658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     19..39
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        21..39
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     enhancer        140..519
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        520..723
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     674..694
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             738..918
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    965..1090
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1583..1816
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2001..2045
                     /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"
     misc_feature    2338..2455
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     regulatory      3572..3581
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     LTR             4243..4476
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4548..4669
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4585..4604)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4639..4658
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4709..4844
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4864..4883)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4865..4883)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4893..4910)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4893..4909)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4902..4924)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5051..5506
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5138..5157)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5348..5369
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5532..5636
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5637..5705,5706..6497)
                     /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(5855..5874)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6668..7256
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7157..7176
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7410..7427
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7544..7565
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7580..7585,7586..7603,7604..7610)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7618..7634
                     /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     7623..7645
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7642..7658
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7642..7658
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 catgattacg ccaagcgcgc aattaaccct cactaaaggg aacaaaagct ggagctgcaa
       61 gcttggccat tgcatacgtt gtatccatat cataatatgt acatttatat tggctcatgt
      121 ccaacattac cgccatgttg acattgatta ttgactagtt attaatagta atcaattacg
      181 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
      241 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
      301 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
      361 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
      421 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
      481 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
      541 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
      601 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
      661 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
      721 gctcgtttag tgaaccgggg tctctctggt tagaccagat ctgagcctgg gagctctctg
      781 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
      841 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
      901 tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga
      961 ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg
     1021 gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg
     1081 cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag
     1141 gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga
     1201 acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg
     1261 acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt
     1321 agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga
     1381 caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct
     1441 tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat aaatataaag
     1501 tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga
     1561 gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa
     1621 gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta
     1681 tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac
     1741 tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa
     1801 aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg
     1861 tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct
     1921 ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag
     1981 aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa
     2041 gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga
     2101 tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag
     2161 ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg
     2221 acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat
     2281 tagtgaacgg atctcgacgg tatcgatctc gacacaaatg gcagtattca tccacaattt
     2341 taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag acataatagc
     2401 aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa attttcgggt
     2461 ttattacagg gacagcagag atccagtttg ggtcgaggaa aaagtggcta agatctacag
     2521 ctgccttgta agtcattggt cttaaaggta ccgagctcga attccgcggg cccgtcgagt
     2581 tggccgcttt atcacttgtt ccgagcccag ttgccgccca ggtggctgta ctgagcatca
     2641 tctcgatctc ggaggggctg atagacctgg tcattcctca acagagcttg tgtgtcggca
     2701 gccccagaca gccttccagt ctcatgtcca gcaaagcaga agactcccaa agcaaggagc
     2761 agagtggcaa tgacatcagt gacaatgatg ccagccacgg tggctggatc cagctccaca
     2821 cagctctggc acactgtggg ggaagggagg agagaggaga ggttgagagc ctttaagatc
     2881 agggaaccat cctctgcctc ctagggcaac ccttagatct cttatgccaa aacccaaact
     2941 tcaataagac cctgggagaa aggcctggtg atgggcttgc cacaccttcc tccaccctgc
     3001 atcctaagga atccctggga agagccaaga agtaatctca gcttttggga ccttgaacaa
     3061 ggtggtgggc caaacctctc acccaaccaa ggctgcagta acatgacccc tactgctctg
     3121 tcttgctgag ttaagctctc tcttccactt tgaaggtccc caaatctggc ttgtaccatt
     3181 acaatagtga cctcactttg tttagagaac agatgggttc accatatccc tctctagcca
     3241 gaaagttctc acatccagaa gccctatcca ttccaaccca aagggttcag gaagcatgta
     3301 cttcgataat gaacttgcac ggtagattct ttgtccttgt atatatctgt cccattacac
     3361 ctatatattc ctcatgggtc caggatgcgt tttcccaggt ccagtcttgt aatgtctgag
     3421 agcagtgttc ccaccgttcc ctctacccat gtgatgctgg tattgcaatt cacaaacact
     3481 ctgtcctcaa gttcctctat aggtatcttg aaggggctca cttgcgagag aagggtagcc
     3541 agtaccaggc cagacaggaa ggtgctgtgc tccatggtgg cgaccggtgg atcccgggcc
     3601 cgcggtaccg tcgactgcag aattcgaagc ttgagctcga ctgatcaccc gggatccccg
     3661 ggtaccgagc tcgaattcat cgatcgcgcc gagtgagggg ttgtgggctc ttttattgag
     3721 ctcggggagc agaagcgcgc gaacagaagc gagaagcgaa ctgattggtt agttcaaata
     3781 aggcacaggg tcatttcagg tccttggggc accctggaaa catctgatgg ttctctagaa
     3841 actgctgagg gcgggaccgc atctggggac catctgttct tggccctgag ccggggcagg
     3901 aactgcttac cacagatatc ctgtttggcc catattctgc tgttccaact gttcttggcc
     3961 ctgagccggg gcaggaactg cttaccacag atatcctgtt tggcccatat tctgctgtct
     4021 ctctgttcct aaccttgatc ctagcttgcc aaacctacag gtggggtctt tcattccccc
     4081 ctttttctgg agactaaata aaatctttta ttctatctat ggctcgtact ctataggctt
     4141 cagctggtga tattgttgag tcaaaactag agcctggacc actgatatcc tgtctttaac
     4201 aaattggact aatcgatctc gatataaaag aaaagggggg actggaaggg ctaattcact
     4261 cccaacgaag acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct
     4321 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     4381 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     4441 tcagaccctt ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta
     4501 ttcagtattt ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg
     4561 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     4621 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctggc
     4681 tctagctatc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     4741 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     4801 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggga cgtacccaat
     4861 tcgccctata gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac
     4921 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     4981 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     5041 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     5101 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     5161 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     5221 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     5281 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     5341 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     5401 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     5461 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     5521 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     5581 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     5641 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     5701 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     5761 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5821 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     5881 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5941 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     6001 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     6061 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     6121 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     6181 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     6241 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6301 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6361 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6421 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6481 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     6541 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     6601 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     6661 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     6721 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     6781 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     6841 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     6901 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     6961 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     7021 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     7081 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     7141 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     7201 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     7261 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     7321 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     7381 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     7441 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     7501 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     7561 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
     7621 tgagcggata acaatttcac acaggaaaca gctatgac
//
