LOCUS       sequence-176104-        6566 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses DsRed in eyes and provides a backbone for expressing two 
            gRNAs. With attB integration site..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6566)
  AUTHORS   Stern DL, Crocker J, Ding Y, Frankel N, Kappes G, Kim E, Kuzmickas 
            R, Lemire A, Mast JD, Picard S
  TITLE     Genetic and Transgenic Reagents for Drosophila simulans, D. 
            mauritiana, D. yakuba, D. santomea, and D. virilis.
  JOURNAL   G3 (Bethesda). 2017 Apr 3;7(4):1339-1347. doi: 
            10.1534/g3.116.038885.
  PUBMED    28280212
REFERENCE   2  (bases 1 to 6566)
  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"; doi: "10.1534/g3.116"; 
            journalName: "G3 (Bethesda)"; date: "2017-04-3- 3"; volume: "7"; 
            issue: "4"; pages: "1339-1347"
COMMENT     Sequence Label: p{CFD4-3xP3::DsRed}
FEATURES             Location/Qualifiers
     source          1..6566
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(1..3,4..6,7..681)
                     /codon_start=1
                     /product="wild-type DsRed"
                     /label=DsRed1
                     /note="mammalian codon-optimized"
                     /translation="MVRSSKNVIKEFMRFKVRMEGTVNGHEFEIEGEGEGRPYEGHNTV
                     KLKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     VVTVTQDSSLQDGCFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     KALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDSKLDITSHNEDYTIVEQYERTEGRH
                     HLFL"
     primer_bind     complement(184..204)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     596..619
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    789..923
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(826..845)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     880..899
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1026..1095
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     protein_bind    1337..1722
                     /label=gypsy insulator
                     /bound_moiety="Su(Hw), CP190, andMod(mdg4)67.2"
                     /note="chromatin insulator from Drosophila"
     promoter        1736..2532
                     /label=dU6-1 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-1 
                     snRNA (Port et al., 2014)"
     misc_RNA        2551..2626
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2627..3047
                     /label=dU6-3 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-3 
                     snRNA (Port et al., 2014)"
     misc_RNA        3066..3141
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(3452..3472)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3452..3470)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3491..3507)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3491..3507)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3504..3526)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3515..3531
                     /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)."
     promoter        complement(join(3539..3545,3546..3563,3564..3569))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3584..3605
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3722..3739)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3893..4481)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3973..3992)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4652..5443,5444..5512))
                     /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     5275..5294
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5513..5617)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5644..6100
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5731..5750)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5941..5962
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6226..6248
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6240..6257
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6241..6257
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6267..6286
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6267..6285
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      join(6561..6566,1..4)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 atggtacgct cctccaagaa cgtcatcaag gagttcatgc gcttcaaggt gcgcatggag
       61 ggcaccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag
      121 ggccacaaca ccgtgaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac
      181 atcctgtccc cccagttcca gtacggctcc aaggtgtacg tgaagcaccc cgccgacatc
      241 cccgactaca agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc
      301 gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg ctgcttcatc
      361 tacaaggtga agttcatcgg cgtgaacttc ccctccgacg gccccgtaat gcagaagaaa
      421 accatgggct gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc
      481 gagatccaca aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagtcc
      541 atctacatgg ccaagaagcc cgtgcagctg cccggctact actacgtgga ctccaagctg
      601 gacatcacct cccacaacga ggactacacc atcgtggagc agtacgagcg caccgagggc
      661 cgccaccacc tgttcctgta gagatcataa tcagccatac cacatttgta gaggttttac
      721 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
      781 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
      841 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
      901 atgtatctta tcatgtctgg atcgatggcg cgcctgatat catcgatctc gaggctgcat
      961 ccaacgcgtt gggagctctc cggatcaatt cggcttcagg taccgtcgac gatgtaggtc
     1021 acggtctcga agccgcggtg cgggtgccag ggcgtgccct tgggctcccc gggcgcgtac
     1081 tccacctcac ccatctggtc catcatgatg aacgggtcga ggtggcggta gttgatcccg
     1141 gcgaacgcgc ggcgcaccgg gaagccctcg ccctcgaaac cgctgggcgc ggtggtcacg
     1201 gtgagcacgg gacgtgcgac ggcgtcggct ggtgcggata cgcggggcag cgtcagcggg
     1261 ttctcgacgg tcacggcggg catgtcgaca agccgaattg atccactaga aggcctaatt
     1321 cggtacacta gttggccacg taataagtgt gcgttgaatt tattcgcaaa aacattgcat
     1381 attttcggca aagtaaaatt ttgttgcata ccttatcaaa aaataagtgc tgcatacttt
     1441 ttagagaaac caaataattt tttattgcat acccgttttt aataaaatac attgcatacc
     1501 ctcttttaat aaaaaatatt gcatactttg acgaaacaaa ttttcgttgc atacccaata
     1561 aaagattatt atattgcata cccgttttta ataaaataca ttgcataccc tcttttaata
     1621 aagaatattg catacgttga cgaaacaaat tttcgttgca tacccaataa aagattatta
     1681 tattgcatac cttttcttgc cataccattt agccgatcaa ttgagatctg aattcatttt
     1741 caacgtcctc gatagtatag tggttagtat ccccgcctgt cacgcgggag accggggttc
     1801 aattccccgt cggggagaat ctgtgattct tttttttttt cttttacttt gttatataaa
     1861 caatttttgt tttaattgaa tctaatttgc cattgctttt aggaatctca ggcatccagc
     1921 aagcgtttgt ccgccgaatc gcccatcagt gaagaagatc ctgtggcggc tacgaaaatc
     1981 tccccggcca tgtcggcctc cacctccagc gaaaaaccca tcagcgagct ggccacctct
     2041 gtgctgaccc accgctttcc agactccacc tcctcacccg gcgaacatgg ccttggacga
     2101 atgcagttgt cgatccgcta cagcgcccag cgtcaaaaac tagacgtgac catacacaaa
     2161 atccagaaga taccacttcg cgatcccagc aatatccccg atccgtatgt taagctgtat
     2221 ctgttgcctg gacgcaccaa ggagtcgaaa cgcaagacga gcgtgatcaa ggacaactgc
     2281 aaccccgtct acgatgcatc ctttgagtac ctgatttcca ttgccgaact caggcagacg
     2341 gaactggagg tgacggtgtg cacccaaaag ggattcctat ccggcggtag tcccatcatt
     2401 ggcatggtag gtacccgaaa gcaacccctt agttacagac acagcgcgta cgtccttcgc
     2461 atccttatga ttcccaagta catattctgc aagagtacag tatatatagg aaagatatcc
     2521 gggtgaactt cggggtcttc gagaagacct gttttagagc tagaaatagc aagttaaaat
     2581 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgctcacct
     2641 gtgattgctc ctactcaaat acaaaaacat caaattttct gtcaataaag catatttatt
     2701 tatatttatt ttacaggaaa gaattccttt taaagtgtat tttaacctat aatgaaaaac
     2761 gattaaaaaa aatacataaa ataattcgaa aatttttgaa tagcccaggt tgataaaaat
     2821 tcatttcata cgttttataa cttatgcccc taagtatttt ttgaccatag tgtttcaatt
     2881 ctacattaat tttacagagt agaatgaaac gccacctact cagccaagag gcgaaaaggt
     2941 tagctcgcca agcagagagg gcgccagtgc tcactacttt ttataattct caacttcttt
     3001 ttccagactc agttcgtata tatagaccta ttttcaattt aacgtcgggg tcttcgagaa
     3061 gacctgtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa
     3121 aaagtggcac cgagtcggtg cttttttgcc tacctggagc ctgagagttg ttcaataaaa
     3181 taaaaatgtt tcgttttttt gctttcgcca gtatttatta tttttcatca atatgtattc
     3241 aatttggtat gtatttagta attgtaatat atagacaatg gttttccgtt gacgtacata
     3301 catctgacgt gtgtttattt agacataata gttatgtttt cacatctttt taatgttcgc
     3361 ttaatgcgta tgcattctag acaattgtgc tcggcaacag tatatttgtg gtgtgccaac
     3421 caacaacctg caggagctcc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     3481 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     3541 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     3601 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     3661 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     3721 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     3781 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     3841 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     3901 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     3961 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     4021 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     4081 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     4141 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     4201 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     4261 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     4321 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     4381 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     4441 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     4501 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     4561 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     4621 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     4681 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     4741 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     4801 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     4861 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     4921 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     4981 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     5041 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     5101 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     5161 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     5221 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     5281 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     5341 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     5401 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     5461 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     5521 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     5581 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     5641 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     5701 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     5761 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     5821 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     5881 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5941 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     6001 tataagggat tttgccgatt tcggcctatt ggttaaaaaa atgagctgat ttaacaaaaa
     6061 tttaacgcga attttaacaa aatattaacg cttacaattt ccattcgcca ttcaggctgc
     6121 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
     6181 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
     6241 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtacag
     6301 gcgcgccggg atctaattca attagagact aattcaatta gagctaattc aattaggatc
     6361 caagcttatc gatttcgaac cctcgaccgc cggagtataa atagaggcgc ttcgtctacg
     6421 gagcgacaat tcaattcaaa caagcaaagt gaacacgtcg ctaagcgaaa gctaagcaaa
     6481 taaacaagcg cagctgaaca agctaaacaa tcggggtacc gctagagtcg acggtaccgc
     6541 gggcccggga tccaccggtc gccacc
//
