LOCUS       sequence-373828-        5427 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination vector for the expression of a fluorescent fusion of 
            N-ter Snap tag and a protein of interest into embryos, by mRNA 
            microinjection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5427)
  TITLE     Lemaire Lab Plasmids 2022
REFERENCE   2  (bases 1 to 5427)
  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-Spe4-Kozak-Snaptag-RfA
FEATURES             Location/Qualifiers
     source          1..5427
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..125
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(126..194,195..986)
                     /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     complement(344..363)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1157..1745
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1646..1665
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1899..1916
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2033..2054
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2069..2074,2075..2092,2093..2099)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2107..2123
                     /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)."
     primer_bind     2112..2134
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2131..2147
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2131..2147
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2168..2186
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     5'UTR           2211..2253
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     CDS             2269..2814
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     protein_bind    2821..2945
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2970..2975,2976..2993,2994..3000)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             3054..3713
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3120..3139)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             4055..4360
                     /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     4239..4258
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4401..4525)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4783..4802)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4784..4802)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4809..4826)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4809..4825)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4818..4840)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(4966..5421)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5103..5124)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5315..5334
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
       61 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
      121 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
      181 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
      241 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
      301 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
      361 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
      421 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
      481 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
      541 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
      601 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
      661 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
      721 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
      781 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
      841 gatctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
      901 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
      961 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     1021 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     1081 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     1141 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     1201 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     1261 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     1321 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     1381 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     1441 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     1501 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     1561 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     1621 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     1681 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     1741 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     1801 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     1861 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     1921 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     1981 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     2041 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     2101 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
     2161 agctcgaaat taaccctcac taaagggaac aaaagctgga gctaagcttg cttgttcttt
     2221 ttgcagaagc tcagaataaa cgctcaactt tggcagatct cagaaaaaat ggacaaagac
     2281 tgcgaaatga agcgcaccac cctggatagc cctctgggca agctggaact gtctgggtgc
     2341 gaacagggcc tgcaccgtat catcttcctg ggcaaaggaa catctgccgc cgacgccgtg
     2401 gaagtgcctg ccccagccgc cgtgctgggc ggaccagagc cactgatgca ggccaccgcc
     2461 tggctcaacg cctactttca ccagcctgag gccatcgagg agttccctgt gccagccctg
     2521 caccacccag tgttccagca ggagagcttt acccgccagg tgctgtggaa actgctgaaa
     2581 gtggtgaagt tcggagaggt catcagctac agccacctgg ccgccctggc cggcaatccc
     2641 gccgccaccg ccgccgtgaa aaccgccctg agcggaaatc ccgtgcccat tctgatcccc
     2701 tgccaccggg tggtgcaggg cgacctggac gtggggggct acgagggcgg gctcgccgtg
     2761 aaagagtggc tgctggccca cgagggccac agactgggca agcctgggct gggtgtaatc
     2821 acaagtttgt acaaaaaagc tgaacgagaa acgtaaaatg atataaatat caatatatta
     2881 aattagattt tgcataaaaa acagactaca taatactgta aaacacaaca tatccagtca
     2941 ctatggcggc cgcattaggc accccaggct ttacacttta tgcttccggc tcgtataatg
     3001 tgtggatttt gagttaggat ccggcgagat tttcaggagc taaggaagct aaaatggaga
     3061 aaaaaatcac tggatatacc accgttgata tatcccaatg gcatcgtaaa gaacattttg
     3121 aggcatttca gtcagttgct caatgtacct ataaccagac cgttcagctg gatattacgg
     3181 cctttttaaa gaccgtaaag aaaaataagc acaagtttta tccggccttt attcacattc
     3241 ttgcccgcct gatgaatgct catccggaat tccgtatggc aatgaaagac ggtgagctgg
     3301 tgatatggga tagtgttcac ccttgttaca ccgttttcca tgagcaaact gaaacgtttt
     3361 catcgctctg gagtgaatac cacgacgatt tccggcagtt tctacacata tattcgcaag
     3421 atgtggcgtg ttacggtgaa aacctggcct atttccctaa agggtttatt gagaatatgt
     3481 ttttcgtctc agccaatccc tgggtgagtt tcaccagttt tgatttaaac gtggccaata
     3541 tggacaactt cttcgccccc gttttcacca tgggcaaata ttatacgcaa ggcgacaagg
     3601 tgctgatgcc gctggcgatt caggttcatc atgccgtctg tgatggcttc catgtcggca
     3661 gaatgcttaa tgaattacaa cagtactgcg atgagtggca gggcggggcg taaacgcgtg
     3721 gatccggctt actaaaagcc agataacagt atgcgtattt gcgcgctgat ttttgcggta
     3781 taagaatata tactgatatg tatacccgaa gtatgtcaaa aagaggtgtg ctatgaagca
     3841 gcgtattaca gtgacagttg acagcgacag ctatcagttg ctcaaggcat atatgatgtc
     3901 aatatctccg gtctggtaag cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa
     3961 cgctggaaag cggaaaatca ggaagggatg gctgaggtcg cccggtttat tgaaatgaac
     4021 ggctcttttg ctgacgagaa cagggactgg tgaaatgcag tttaaggttt acacctataa
     4081 aagagagagc cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgcccgg
     4141 gcgacggatg gtgatccccc tggccagtgc acgtctgctg tcagataaag tctcccgtga
     4201 actttacccg gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc
     4261 cagtgtgccg gtctccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga
     4321 catcaaaaac gccattaacc tgatgttctg gggaatataa atgtcaggct cccttataca
     4381 cagccagtct gcaggtcgac catagtgact ggatatgttg tgttttacag tattatgtag
     4441 tctgtttttt atgcaaaatc taatttaata tattgatatt tatatcattt tacgtttctc
     4501 gttcagcttt cttgtacaaa gtggtgataa acttagatat cgcggccgcg gaagcgctgg
     4561 ttacctagga accagcctca agaacacccg aatggagtct ctaagctaca taataccaac
     4621 ttacacttta caaaatgttg tcccccaaaa tgtagccatt cgtatctscy ccwaawaaaa
     4681 araaarttty ttcmcawtcw aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaamcccccc
     4741 cccccccccc ctgcagggcc aagtcggccg gcgcgccatt cgccctatag tgagtcgtat
     4801 tacaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     4861 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     4921 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggaaatt gtaagcgtta
     4981 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     5041 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     5101 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     5161 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
     5221 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     5281 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     5341 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     5401 atgcgccgct acagggcgcg tcaggtg
//
