LOCUS       sequence-169409-        6446 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 6446)
  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 6446)
  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..6446
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     170..193
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    363..497
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(400..419)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     454..473
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    600..669
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     promoter        1194..1990
                     /label=dU6-1 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-1 
                     snRNA (Port et al., 2014)"
     misc_RNA        2009..2084
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2085..2505
                     /label=dU6-3 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-3 
                     snRNA (Port et al., 2014)"
     misc_RNA        2524..2599
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(2910..2930)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2910..2928)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2949..2965)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2949..2965)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2962..2984)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2973..2989
                     /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(2997..3003,3004..3021,3022..3027))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3042..3063
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3180..3197)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3351..3939)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3431..3450)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4110..4901,4902..4970))
                     /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     4733..4752
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4971..5075)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5101..5556)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5238..5259)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5450..5469
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5683..5705
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5697..5714
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5698..5714
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5721..5740
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5721..5739
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      6015..6024
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(6021..6023,6024..6026,6027..6446,1..255)
                     /codon_start=1
                     /product="wild-type DsRed"
                     /label=DsRed1
                     /note="mammalian codon-optimized"
                     /translation="MVRSSKNVIKEFMRFKVRMEGTVNGHEFEIEGEGEGRPYEGHNTV
                     KLKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     VVTVTQDSSLQDGCFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     KALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDSKLDITSHNEDYTIVEQYERTEGRH
                     HLFL"
     primer_bind     complement(6204..6224)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
ORIGIN
        1 ggctgggagg cctccaccga gcgcctgtac ccccgcgacg gcgtgctgaa gggcgagatc
       61 cacaaggccc tgaagctgaa ggacggcggc cactacctgg tggagttcaa gtccatctac
      121 atggccaaga agcccgtgca gctgcccggc tactactacg tggactccaa gctggacatc
      181 acctcccaca acgaggacta caccatcgtg gagcagtacg agcgcaccga gggccgccac
      241 cacctgttcc tgtagagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
      301 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
      361 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
      421 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
      481 cttatcatgt ctggatcgat ggcgcgcctg atatcatcga tctcgaggct gcatccaacg
      541 cgttgggagc tctccggatc aattcggctt caggtaccgt cgacgatgta ggtcacggtc
      601 tcgaagccgc ggtgcgggtg ccagggcgtg cccttgggct ccccgggcgc gtactccacc
      661 tcacccatct ggtccatcat gatgaacggg tcgaggtggc ggtagttgat cccggcgaac
      721 gcgcggcgca ccgggaagcc ctcgccctcg aaaccgctgg gcgcggtggt cacggtgagc
      781 acgggacgtg cgacggcgtc ggctgggtgc ggatacgcgg ggcagcgtca gcgggttctc
      841 gacggtcacg gcgggcatgt cgacaagccg aattgatcca ctagaaggcc taattcggta
      901 cactagttgg ccacgtaata agtgtgcgtt gaatttattc gcaaaaacat tgcatatttt
      961 cggcaaagta aaattttgtt gcatacctta tcaaaaaata agtgctgcat actttttaga
     1021 gaaaccaaat aattttttat tgcatacccg tttttaataa aatacattgc ataccctctt
     1081 ttaataaaaa atattgcata ctttgacgaa acaaattttc gttgcatacc caataaaaga
     1141 ttattatatt gcatacccgt ttttaataaa atacattgca tagatctgaa ttcattttca
     1201 acgtcctcga tagtatagtg gttagtatcc ccgcctgtca cgcgggagac cggggttcaa
     1261 ttccccgtcg gggagaatct gtgattcttt ttttttttct tttactttgt tatataaaca
     1321 atttttgttt taattgaatc taatttgcca ttgcttttag gaatctcagg catccagcaa
     1381 gcgtttgtcc gccgaatcgc ccatcagtga agaagatcct gtggcggcta cgaaaatctc
     1441 cccggccatg tcggcctcca cctccagcga aaaacccatc agcgagctgg ccacctctgt
     1501 gctgacccac cgctttccag actccacctc ctcacccggc gaacatggcc ttggacgaat
     1561 gcagttgtcg atccgctaca gcgcccagcg tcaaaaacta gacgtgacca tacacaaaat
     1621 ccagaagata ccacttcgcg atcccagcaa tatccccgat ccgtatgtta agctgtatct
     1681 gttgcctgga cgcaccaagg agtcgaaacg caagacgagc gtgatcaagg acaactgcaa
     1741 ccccgtctac gatgcatcct ttgagtacct gatttccatt gccgaactca ggcagacgga
     1801 actggaggtg acggtgtgca cccaaaaggg attcctatcc ggcggtagtc ccatcattgg
     1861 catggtaggt acccgaaagc aaccccttag ttacagacac agcgcgtacg tccttcgcat
     1921 ccttatgatt cccaagtaca tattctgcaa gagtacagta tatataggaa agatatccgg
     1981 gtgaacttcg gggtcttcga gaagacctgt tttagagcta gaaatagcaa gttaaaataa
     2041 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt gctcacctgt
     2101 gattgctcct actcaaatac aaaaacatca aattttctgt caataaagca tatttattta
     2161 tatttatttt acaggaaaga attcctttta aagtgtattt taacctataa tgaaaaacga
     2221 ttaaaaaaaa tacataaaat aattcgaaaa tttttgaata gcccaggttg ataaaaattc
     2281 atttcatacg ttttataact tatgccccta agtatttttt gaccatagtg tttcaattct
     2341 acattaattt tacagagtag aatgaaacgc cacctactca gccaagaggc gaaaaggtta
     2401 gctcgccaag cagagagggc gccagtgctc actacttttt ataattctca acttcttttt
     2461 ccagactcag ttcgtatata tagacctatt ttcaatttaa cgtcggggtc ttcgagaaga
     2521 cctgttttag agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa
     2581 agtggcaccg agtcggtgct tttttgccta cctggagcct gagagttgtt caataaaata
     2641 aaaatgtttc gtttttttgc tttcgccagt atttattatt tttcatcaat atgtattcaa
     2701 tttggtatgt atttagtaat tgtaatatat agacaatggt tttccgttga cgtacataca
     2761 tctgacgtgt gtttatttag acataatagt tatgttttca catcttttta atgttcgctt
     2821 aatgcgtatg cattctagac aattgtgctc ggcaacagta tatttgtggt gtgccaacca
     2881 acaacctgca ggagctccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg
     2941 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     3001 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3061 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     3121 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3181 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3241 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3301 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3361 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3421 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3481 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3541 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3601 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3661 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3721 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3781 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3841 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3901 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3961 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4021 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4081 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4141 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4201 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4261 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4321 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4381 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4441 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4501 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4561 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4621 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4681 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4741 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4801 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4861 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4921 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4981 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5041 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5101 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     5161 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     5221 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     5281 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     5341 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     5401 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     5461 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     5521 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
     5581 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
     5641 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
     5701 aaacgacggc cagtgaattg taatacgact cactataggg cgaattgggt acaggcgcgc
     5761 cgggatctaa ttcaattaga gactaattca attagagcta attcaattag gatccaagct
     5821 tatcgatttc gaaccctcga ccgccggagt ataaatagag gcgcttcgtc tacggagcga
     5881 caattcaatt caaacaagca aagtgaacac gtcgctaagc gaaagctaag caaataaaca
     5941 agcgcagctg aacaagctaa acaatcgggg taccgctaga gtcgacggta ccgcgggccc
     6001 gggatccacc ggtcgccacc atggtgcgct cctccaagaa cgtcatcaag gagttcatgc
     6061 gcttcaaggt gcgcatggag ggcaccgtga acggccacga gttcgagatc gagggcgagg
     6121 gcgagggccg cccctacgag ggccacaaca ccgtgaagct gaaggtgacc aagggcggcc
     6181 ccctgccctt cgcctgggac atcctgtccc cccagttcca gtacggctcc aaggtgtacg
     6241 tgaagcaccc cgccgacatc cccgactaca agaagctgtc cttccccgag ggcttcaagt
     6301 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc
     6361 tgcaggacgg ctgcttcatc tacaaggtga agttcatcgg cgtgaacttc ccctccgacg
     6421 gccccgtaat gcagaagaaa accatg
//
