LOCUS       sequence-348367-        6374 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for generating PCR templates with sfGFP and Amp for BAC 
            recombination.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6374)
  AUTHORS   Shih YH, Portman D, Idrizi F, Grosse A, Lawson ND
  TITLE     Integrated molecular analysis identifies a conserved pericyte gene 
            signature in zebrafish.
  JOURNAL   Development. 2021 Nov 9. pii: 273393. doi: 10.1242/dev.200189.
  PUBMED    34751773
REFERENCE   2  (bases 1 to 6374)
  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: 
            "Development. 2021 Nov 9. pii: 273393. doi: 10.1242/dev.200189."
COMMENT     Sequence Label: p-sfGFP-pA-FRT-Amp-FRT
FEATURES             Location/Qualifiers
     source          1..6374
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        108..411
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        412..615
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     566..586
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     polyA_signal    1368..1502
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1564..1582
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1564..1581
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    1657..1690
                     /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)"
     CDS             complement(join(1845..2636,2637..2705))
                     /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     2468..2487
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2706..2810)
                     /gene="bla"
                     /label=AmpR promoter
     protein_bind    2968..3001
                     /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)"
     regulatory      3012..3021
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3051..3070)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3052..3070)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    3209..3330
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(3337..3792)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3474..3495)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3686..3705
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3819..3923
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3925..4282
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3925..4116
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3948..3968)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4133..4268
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4317..5111
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4317..5111
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4371..4390)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4981..5000
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5299..5318)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5343..5390
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5719..6307
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6208..6227
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcattag ttattaatag
       61 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
      121 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
      181 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
      241 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
      301 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
      361 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      421 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      481 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      541 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
      601 tatataagca gagctggttt agtgaaccgt cagatccggc caccatgtcc aaaggagaag
      661 agctgttcac tggagtggta ccaatacttg tggagttgga cggagatgtg aacggacaca
      721 aattttcagt ccgcggggag ggggaagggg atgctactat tggcaagctg acgctcaaat
      781 tcatctgtac caccggaaaa ctccctgtac cctggcccac actggtgaca actctgactt
      841 acggcgtgca atgttttagc cgatacccag accacatgaa gaggcacgac tttttcaaaa
      901 gcgcaatgcc tgaaggatac gtacaggaaa ggaccatttc ttttaaagac gacgggaagt
      961 acaaaacccg ggcagtggtg aagtttgagg gcgataccct cgtcaatagg atcgaattga
     1021 agggaactga cttcaaagaa gatggcaaca tcctgggtca caagcttgag tataacttta
     1081 actcccacaa cgtgtatatt acagccgaca aacagaagaa tggaattaag gctaacttca
     1141 ctgtcagaca caatgtcgaa gatggctccg tgcagctcgc cgatcactat caacagaata
     1201 ctcctatcgg ggacggccca gtcctgctgc ccgacaacca ctacctgagt acccagactg
     1261 ttctgagcaa agatccgaac gagaagcgcg accgcatggt gttgcatgag tatgtcaacg
     1321 ctgcgggaat taccctcggc atggacgagc tgtacaagtc tagataggat ccagacatga
     1381 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     1441 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     1501 ttaacaacaa caattgcatt cactttatgt ttcaggttca gggggaggtg tgggaggttt
     1561 tttatttagg tgacactata gaatactcaa gctatgcatc caacgcgttg ggagctctcc
     1621 catatggtcg acctgcaggc ggccgcacta gtgattgaag ttcctattct ctagaaagta
     1681 taggaacttc tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     1741 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     1801 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     1861 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     1921 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     1981 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     2041 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     2101 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     2161 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     2221 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     2281 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     2341 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     2401 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     2461 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     2521 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     2581 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     2641 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     2701 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     2761 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     2821 cgaaaagtgc cacctgatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     2881 catcaggaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     2941 agctcatttt ttaaccaata ggccgaagaa gttcctattc tctagaaagt ataggaactt
     3001 caatcccgcg gccatggcgg ccgggagcat gcgacgtcgg gcccaattcg ccctatagtg
     3061 agtcgtatta ccgggcccgg gatccaccgg atctagataa ctgatcataa tcagccatac
     3121 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     3181 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     3241 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     3301 tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt aatattttgt
     3361 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     3421 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     3481 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     3541 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     3601 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     3661 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     3721 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     3781 tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     3841 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     3901 aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca
     3961 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4021 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4081 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4141 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4201 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4261 tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga
     4321 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     4381 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     4441 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag
     4501 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     4561 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     4621 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     4681 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4741 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     4801 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg
     4861 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4921 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     4981 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     5041 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     5101 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     5161 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     5221 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     5281 ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat
     5341 gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg
     5401 ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata
     5461 cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct
     5521 cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat atatacttta
     5581 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     5641 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     5701 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5761 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5821 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     5881 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     5941 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     6001 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     6061 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     6121 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     6181 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     6241 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     6301 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     6361 tcacatgttc tttc
//
