LOCUS       sequence-358577-        3666 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For generation of plasmids for CRISPR-mediated 3p3 GFP knockin in 
            Aedes aegypti.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3666)
  AUTHORS   Chen J, Luo J, Gurav AS, Chen Z, Wang Y, Montell C
  TITLE     A DREaMR system to simplify combining mutations with rescue 
            transgenes in Aedes aegypti.
  JOURNAL   Genetics. 2021 Nov 5;219(3). pii: 6368066. doi: 
            10.1093/genetics/iyab146.
  PUBMED    34740249
REFERENCE   2  (bases 1 to 3666)
  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: "Genetics.";
            date: "2021-11-5"; volume: "219(3). pii: 6368066. doi"; pages: " 
            10.1093/genetics/iyab146"
COMMENT     Sequence Label: AeU6 3p3 GFP LgRNA
FEATURES             Location/Qualifiers
     source          1..3666
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(80..871,872..940))
                     /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     703..722
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(941..1045)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      1344..1353
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1350..1352,1353..1355,1356..2069)
                     /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(1350..1352,1353..1355,1356..2066)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1395..1416)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1656..1675)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2003..2024
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2210..2331
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2247..2266)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2596..2612)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2596..2612)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    2831..2900
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     rep_origin      complement(2987..3575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3067..3086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
       61 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
      121 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
      181 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
      241 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
      301 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
      361 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
      421 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
      481 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
      541 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
      601 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
      661 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
      721 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
      781 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
      841 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
      901 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
      961 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     1021 aaaataaaca aataggggtt ccgcgcacat ttccttaatt aactcgcgtt aagatacatt
     1081 gatgagtttg gacaaacctg caggcccttt cgtcttcaag aattggatct aattcaatta
     1141 gagactaatt caattagagc taattcaatt aggatccaag cttatcgatt tcgaaccctc
     1201 gaccgccgga gtataaatag aggcgcttcg tctacggagc gacaattcaa ttcaaacaag
     1261 caaagtgaac acgtcgctaa gcgaaagcta agcaaataaa caagcgcagc tgaacaagct
     1321 aaacaatcgg ctcgaagccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg
     1381 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
     1441 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
     1501 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt
     1561 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
     1621 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
     1681 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
     1741 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
     1801 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
     1861 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
     1921 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag
     1981 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
     2041 ctctcggcat ggacgagctg tacaagtaaa gcggcctaag cgatcgctca ggccgcgact
     2101 ctagatcata atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc
     2161 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
     2221 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     2281 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt acctgcagga
     2341 gtaaactcgt cgacggtatc gataagcgct agcgaatgaa atcgcccatc gagttgatac
     2401 gtccatccat cgctagaacc gcgttcgctg tagaagacta tataagagca gaggcaagag
     2461 tagtgaaatg gtaccggccg cgtcttagag ctatgctgga aacagcatag caagttaaga
     2521 taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt tttatcgaat
     2581 tcctgcagcc cgggggatcc actagttcta gagcggcgcg cccatatgtt cggcttgtcg
     2641 acatgcccgc cgtgaccgtc gagaacccgc tgacgctgcc ccgcgtatcc gcacccgccg
     2701 acgccgtcgc acgtcccgtg ctcaccgtga ccaccgcgcc cagcggtttc gagggcgagg
     2761 gcttcccggt gcgccgcgcg ttcgccggga tcaactaccg ccacctcgac ccgttcatca
     2821 tgatggacca gatgggtgag gtggagtacg cgcccgggga gcccaagggc acgccctggc
     2881 acccgcaccg cggcttcgag accgtgacct acatcgtcga cggtaacatg tgagcaaaag
     2941 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     3001 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3061 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3121 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     3181 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     3241 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     3301 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     3361 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     3421 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     3481 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     3541 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     3601 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     3661 aaagga
//
