LOCUS       sequence-13966-e        4955 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4955)
  TITLE     Sekelsky lab Drosophila phiC31 cloning plasmids
REFERENCE   2  (bases 1 to 4955)
  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: pDEST-attB
FEATURES             Location/Qualifiers
     source          1..4955
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(149..154,155..172,173..179)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             233..892
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(299..318)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1234..1539
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     1418..1437
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(1580..1704)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     1727..1743
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1728..1744
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1781..1797)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1781..1797)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    2015..2084
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     primer_bind     complement(2121..2140)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2122..2140)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2159..2175)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2159..2175)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2172..2194)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2183..2199
                     /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(2207..2213,2214..2231,2232..2237))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2252..2273
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2390..2407)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2561..3149)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2641..2660)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3320..4111,4112..4180))
                     /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     3943..3962
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4181..4285)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4312..4767
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4399..4418)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4609..4630
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4894..4916
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4908..4925
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4909..4925
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    join(4955,1..24)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
ORIGIN
        1 caagtttgta caaaaaagct gaacgagaaa cgtaaaatga tataaatatc aatatattaa
       61 attagatttt gcataaaaaa cagactacat aatactgtaa aacacaacat atccagtcac
      121 tatggcggcc gcattaggca ccccaggctt tacactttat gcttccggct cgtataatgt
      181 gtggattttg agttaggatc cgtcgagatt ttcaggagct aaggaagcta aaatggagaa
      241 aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga
      301 ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc
      361 ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct
      421 tgcccgcctg atgaatgctc atccggaatt ccgtatggca atgaaagacg gtgagctggt
      481 gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc
      541 atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga
      601 tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt
      661 tttcgtctca gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat
      721 ggacaacttc ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt
      781 gctgatgccg ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag
      841 aatgcttaat gaattacaac agtactgcga tgagtggcag ggcggggcgt aaagatctgg
      901 atccggctta ctaaaagcca gataacagta tgcgtatttg cgcgctgatt tttgcggtat
      961 aagaatatat actgatatgt atacccgaag tatgtcaaaa agaggtatgc tatgaagcag
     1021 cgtattacag tgacagttga cagcgacagc tatcagttgc tcaaggcata tatgatgtca
     1081 atatctccgg tctggtaagc acaaccatgc agaatgaagc ccgtcgtctg cgtgccgaac
     1141 gctggaaagc ggaaaatcag gaagggatgg ctgaggtcgc ccggtttatt gaaatgaacg
     1201 gctcttttgc tgacgagaac aggggctggt gaaatgcagt ttaaggttta cacctataaa
     1261 agagagagcc gttatcgtct gtttgtggat gtacagagtg atattattga cacgcccggg
     1321 cgacggatgg tgatccccct ggccagtgca cgtctgctgt cagataaagt ctcccgtgaa
     1381 ctttacccgg tggtgcatat cggggatgaa agctggcgca tgatgaccac cgatatggcc
     1441 agtgtgccgg tctccgttat cggggaagaa gtggctgatc tcagccaccg cgaaaatgac
     1501 atcaaaaacg ccattaacct gatgttctgg ggaatataaa tgtcaggctc ccttatacac
     1561 agccagtctg caggtcgacc atagtgactg gatatgttgt gttttacagt attatgtagt
     1621 ctgtttttta tgcaaaatct aatttaatat attgatattt atatcatttt acgtttctcg
     1681 ttcagctttc ttgtacaaag tggttgatgg ggatccgaat tcgagctcga ggtcgacggt
     1741 atcgataagc ttgatatcga attcctgcag cccgggcagg gatccactag ttctagaatt
     1801 cgagctcccg ggtactctag gtcgacatgc ccgccgtgac cgtcgagaac ccgctgacgc
     1861 tgccccgcgt atccgcaccc gccgacgccg tcgcacgtcc cgtgctcacc gtgaccaccg
     1921 cgcccagcgg tttcgagggc gagggcttcc cggtgcgccg cgcgttcgcc gggatcaact
     1981 accgccacct cgacccgttc atcatgatgg accagatggg tgaggtggag tacgcgcccg
     2041 gggagcccaa gggcacgccc tggcacccgc accgcggctt cgagaccgtg acctacatcg
     2101 tcgacgcggc cacaagcttt ccctatagtg agtcgtatta gagcttggcg taatcatggt
     2161 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     2221 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     2281 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     2341 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     2401 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     2461 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     2521 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     2581 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     2641 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     2701 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     2761 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     2821 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     2881 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     2941 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3001 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3061 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3121 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3181 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     3241 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     3301 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     3361 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     3421 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     3481 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     3541 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     3601 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     3661 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     3721 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     3781 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     3841 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     3901 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     3961 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     4021 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     4081 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     4141 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     4201 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4261 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgcgccct
     4321 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     4381 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     4441 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     4501 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     4561 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     4621 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     4681 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     4741 ttaacaaaat attaacgttt acaatttccc attcgccatt caggctgcgc aactgttggg
     4801 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg
     4861 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg
     4921 ccagtgaatt cgagctcggt accgggcccc atcaa
//
