LOCUS       sequence-210555-        4979 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recipient vector for Cranial Neural Crest-specific expression of 
            protein of interest.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4979)
  TITLE     Cranial Neural Crest-specific expression of protein of interest
REFERENCE   2  (bases 1 to 4979)
  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: pTK-enhNC1.1_M3-MCS-2A-eGFP-SV40pA
FEATURES             Location/Qualifiers
     source          1..4979
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..49
                     /note="synthetic polyadenylation signal"
     misc_feature    63..154
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     103..122
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        834..979
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          1057..1189
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1172..1191
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1247..1256
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1272..1325
                     /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"
     CDS             join(1332..1334,1335..1337,1338..2051)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1332..1334,1335..1337,1338..2048)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1377..1398)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1638..1657)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1985..2006
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2103..2224
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2114..2133)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2168..2187
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2472..2489)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2643..3231)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2723..2742)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3402..4193,4194..4262))
                     /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     4025..4044
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4263..4367)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4394..4849
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4481..4500)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4691..4712
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgatagta
       61 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      121 cccagtgcaa gtgcaggtgc cagaacattt ctctatcgat aggtacccag taagctttcc
      181 accaacactt aacctttatg aagaaccaaa ttattttgtt ccatgactaa gaaaaatctg
      241 gcgatatata attaaaacaa caaagaagag tcccattcta tacaagtgta attacgttca
      301 tgtggtatca ctgtaattga aacacagaag atgtttcagt ctgtattaaa agagaatttc
      361 tttcagtggg tttcctgaag ctcattagat attccctggc ctattctcct aagtaattag
      421 attttaacag gattttcaac agatcagagc aatctctgtg ggtgccaata ggagactctc
      481 cctctggcct taccagcagg gcagcatgga taacatccag gctgggagtt aataggacaa
      541 ttactggcct cttttaaatg agttacatta atcgtggagt acaactacaa cagccacaac
      601 gtctatatca tggccgacaa gcagaagaac ggcatcgagg tgaacttcaa gatccgccac
      661 aacatcgagg acggcggatt ttggtggtgt atgtaaatgt gcattggata gtgtatgtta
      721 aacctcgaga tctgcgatct aagtaagctt gctgcttcat ccccgtggcc cgttgctcgc
      781 gtttgctggc ggtgtccccg gaagaaatat atttgcatgt ctttagttct atgatgacac
      841 aaaccccgcc cagcgtcttg tcattggcga attcgaacac gcagatgcag tcggggcggc
      901 gcggtcccag gtccacttcg catattaagg tgacgcgtgt ggcctcgaac accgagcgac
      961 cctgcagcga cccgcttaac agcgtcaaca gcgtgccgca gatcccaagc ttgcggccgc
     1021 ttaagctgca gaagttggtc gtgaggcact gggcaggtaa gtatcaaggt tacaagacag
     1081 gtttaaggag accaatagaa actgggcttg tcgagacaga gaagactctt gcgtttctga
     1141 taggcaccta ttggtcttac tgacatccac tttgcctttc tctccacagg tgtccactcc
     1201 caggttcaat acagctctta agcggccgca agctagctac gtagccgcca ccatgggata
     1261 tcggatcagg agagggcagg ggaagtcttc taacatgcgg ggacgtggag gaaaatcccg
     1321 ggcccagatc tatggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     1381 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     1441 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     1501 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     1561 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     1621 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     1681 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     1741 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     1801 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     1861 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     1921 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     1981 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     2041 tgtacaagta aagcggccgc gactctagag tcggggcggc cggccgcttc gagcagacat
     2101 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
     2161 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
     2221 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt
     2281 tttttaaagc aagtaaaacc tctacaaatg tggtaaaatc gataaggatc cgtcgaccga
     2341 tgcccttgag agccttcaac ccagtcagct ccttccggtg ggcgcggggc atgactatcg
     2401 tcgccgcact tatgactgtc ttctttatca tgcaactcgt aggacaggtg ccggcagcgc
     2461 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     2521 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     2581 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     2641 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     2701 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     2761 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     2821 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     2881 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     2941 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     3001 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     3061 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     3121 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     3181 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     3241 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     3301 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     3361 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     3421 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     3481 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     3541 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     3601 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     3661 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     3721 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     3781 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     3841 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     3901 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     3961 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     4021 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     4081 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     4141 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     4201 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     4261 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     4321 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     4381 aaagtgccac ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     4441 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     4501 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     4561 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     4621 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     4681 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     4741 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     4801 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaatttg ccattcgcca
     4861 ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag
     4921 cccaagctac catgataagt aagtaatatt aaggtacggg aggtacttgg agcggccgc
//
