LOCUS Exported 5194 bp ds-DNA circular SYN 28-OCT-2021 DEFINITION Leishmania cell free expression of zebrafish mCherry-Bin1b. Parton lab clone GSU. ACCESSION . VERSION . KEYWORDS SITS-mCherry-Bin1b SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5194) TITLE Harriet's Cavin4 paper JOURNAL Unpublished REFERENCE 2 (bases 1 to 5194) AUTHORS . TITLE Direct Submission JOURNAL Exported Oct 28, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5194 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind 117..137 /gene="mutant version of attB" /label=attB4 /bound_moiety="BP Clonase(TM)" /note="core recombination site for the Gateway(R) BP reaction" primer_bind 168..187 /label=T7 /note="T7 promoter, forward primer" promoter 168..186 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 325..348 /codon_start=1 /product="8xHis affinity tag" /label=8xHis /translation="HHHHHHHH" protein_bind 348..372 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" regulatory 373..382 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 379..1086 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(524..542) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 792..811 /label=mCherry-F /note="mCherry, forward primer" primer_bind 986..1009 /label=DsRed1-C /note="DsRed1, forward primer" protein_bind complement(1088..1112) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" protein_bind complement(2206..2226) /gene="mutant version of attB" /label=attB3 /bound_moiety="BP Clonase(TM)" /note="core recombination site for the Gateway(R) BP reaction" primer_bind complement(2234..2250) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" polyA_signal 2266..2400 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2290..2309) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2344..2363 /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind complement(2565..2585) /label=T3 /note="T3 promoter, forward primer" promoter complement(2565..2583) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(2604..2620) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2604..2620) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(2617..2639) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 2628..2644 /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(2652..2682) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 2697..2718 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(2835..2852) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3006..3594) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3086..3105) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(3765..4625) /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 4388..4407 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(4626..4730) /gene="bla" /label=AmpR promoter rep_origin complement(4756..17) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4893..4914) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 5105..5124 /label=F1ori-R /note="F1 origin, reverse primer" ORIGIN 1 gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg aagggcgatc 61 ggtgcgggcc tcttcgctat tacgccagtc gactagggat aacagggtaa tgaattcaac 121 tttgtataga aaagttgaca tttccccgaa aagtgccacc tgacgtctaa tacgactcac 181 tatagggaca tcttaagttt attttatttt attttatttt attttatttt attttatttt 241 attttatttt atttaaccat gacagtaatg tataaagtct gtaaagacat taaacacgta 301 agtgaaacca tgggcacctc tggtcaccac catcaccacc atcaccacca agtttgtaca 361 aaaaagcagg ctgccaccat ggtgagcaag ggcgaggagg acaacatggc catcatcaag 421 gagttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc 481 gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc 541 aagggcggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc 601 aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag 661 ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag 721 gactcctccc tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc 781 ccctccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc ctccgagcgg 841 atgtaccccg aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac 901 ggcggccact acgacgccga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg 961 cccggcgcct acaacgtcaa catcaagctg gacatcacct cccacaacga ggactacacc 1021 atcgtggaac agtacgagcg cgccgagggc cgccactcca ccggcggcat ggacgagctg 1081 tacaagcacc cagctttctt gtacaaagtg gtggcagagg ttgggaaagg agtcaacgcc 1141 gggaaactgg ccagtaatgt acagaagagg ttaacccgag cacaagagaa ggtgatgcag 1201 aaactgggaa aggcggatga aaccaaagac acagcctttg aggagcaagt agccaacttt 1261 aacaaacaga tgaatgaggg caccaaactg cagaaagact tgaaagcgta tttgctggca 1321 gtcaaaacaa tgcacgagtg ttccaagcgt ctgcacgact gtctggcaga aatgtatgac 1381 cctgagtggt ttggcaaaga ggaggtggac tccatcgcag aggacactga cctcctgtgg 1441 caagatttcc acgagaagct tttggacaat gctgtaattt ccatggacgc atacttggct 1501 cagtttccag atataaaggc tcgcattgca aagcgtgaaa ggaagctggt ggactttgac 1561 agtgccagac atcattttgc ctctgtacaa aaaggcaaga aaaaggacga ggccaaaatt 1621 gccaaggctg aggaacactt gggcagagct cagaaagtgt ttgaggagat caattatgat 1681 ctccaggagg aacttcccac cctgtggaac agtcgtattg gcctctatgt taacacattc 1741 cggagtgtgt ctggcctgga ggaaacattt cacagggaca tgggcaaact gaaccagaat 1801 ctgagtgata ttatgaccaa actggaagag cagcgtgttg ccaaaaaagg tgtcacgaca 1861 gcaagcactg ggaagagtga cgatgcagca aaccacaatc aggagacgtc agatgcacct 1921 aaatctccat caaagaatga agccccaacg gtgacaaatg gctcctcaga tacagagctt 1981 cctccatctg ttctctttaa ggtgaaaaca atgcacgact acagtgcaac agacagtgat 2041 gaacttgacc tgaaggctgg tgacattgtg cttgtgcttc cctttgccaa cccagatgaa 2101 gcggatgatg gctggttaat gggtgtaaaa gagtcccact ggcttcagaa caaaaacctc 2161 ctagcaaaag gagtcttccc tgaaaacttc acccagaaac tgtgacaact ttattataca 2221 tagttgataa ttcactggcc gtcgttttac ggtaccatcg atgatgatcc agacatgata 2281 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt 2341 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt 2401 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt 2461 taaagcaagt aaaacctcta caaatgtggt atggctgatt atgatctcga gattaccctg 2521 ttatccctag cgtggatcct aggactagta cccagctttt gttcccttta gtgagggtta 2581 attgcgcgct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc 2641 acaattccac acaacatacg agccgggagc ataaagtgta aagcctgggg tgcctaatga 2701 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg 2761 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg 2821 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg 2881 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga 2941 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg 3001 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag 3061 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc 3121 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg 3181 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt 3241 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc 3301 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc 3361 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg 3421 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca 3481 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc 3541 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat 3601 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt 3661 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt 3721 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc 3781 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc 3841 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata 3901 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg 3961 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc 4021 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct 4081 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa 4141 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt 4201 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca 4261 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac 4321 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca 4381 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt 4441 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc 4501 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca 4561 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata 4621 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc 4681 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc 4741 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt 4801 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa 4861 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa 4921 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac 4981 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga 5041 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa 5101 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc 5161 tgcgcgtaac caccacaccc gccgcgctta atgc //