LOCUS       Exported                6702 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Mammalian expression of hArl8 with C-term GFP tag.
ACCESSION   .
VERSION     .
KEYWORDS    pDEST47-ARL8A-GFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6702)
  TITLE     Unpublished Kahn lab ARF plasmids
REFERENCE   2  (bases 1 to 6702)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6702
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    680..704
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1281..1305)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1316..2035
                     /codon_start=1
                     /product="Cycle 3 GFP (Crameri et al., 1996)"
                     /label=Cycle 3 GFP
                     /note="mammalian codon-optimized"
                     /translation="MASKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(1349..1377)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1972..1992
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(2057..2074)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2063..2287
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2333..2761
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2420..2439)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2630..2651
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2770..2790)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2775..3104
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2955..3090
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3017..3036
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3171..3965
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3225..3244)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3835..3854
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4141..4262
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4178..4197)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4232..4251
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4311..4327)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4311..4327)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4324..4346)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4335..4351
                     /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(4359..4389)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4404..4425
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4542..4559)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4713..5301)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4793..4812)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5472..6332)
                     /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     6095..6114
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6333..6437)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6512..6531)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagttaag cttgatcaaa caagtttgta caaaaaagca ggcttagaag gagatagaac
      721 catgatcgct ttgttcaaca agctgctgga ctggttcaag gccctattct ggaaggagga
      781 gatggagctc acgctggtcg ggcttcagta ctcgggcaag accaccttcg tcaacgtgat
      841 cgcgtcagga cagttcaacg aggacatgat ccccaccgtg ggtttcaaca tgcgcaaaat
      901 caccaaaggg aatgtgacta tcaagctctg ggacattggg ggacagccgc gtttccgcag
      961 catgtgggag cgctactgcc gaggagtgag cgccatcgtg tacatggtgg atgctgctga
     1021 ccaggagaag attgaggcct ctaagaacga gctccacaac ctactggaca aacctcagct
     1081 gcagggcatc ccggtcttag tcctgggtaa caagcgagac cttccgggag cattggatga
     1141 gaaggagctg attgagaaaa tgaatctgtc tgccatccag gaccgagaga tctgctgcta
     1201 ctccatctct tgcaaagaaa aggacaacat tgacatcacc ctacagtggc ttattcaaca
     1261 ctcgaagtca cggagaagcg acccagcttt cttgtacaaa gtggttcgat ctagaatggc
     1321 tagcaaagga gaagaacttt tcactggagt tgtcccaatt cttgttgaat tagatggtga
     1381 tgttaatggg cacaaatttt ctgtcagtgg agagggtgaa ggtgatgcta catacggaaa
     1441 gcttaccctt aaatttattt gcactactgg aaaactacct gttccatggc caacacttgt
     1501 cactactttc tcttatggtg ttcaatgctt ttcccgttat ccggatcata tgaaacggca
     1561 tgactttttc aagagtgcca tgcccgaagg ttatgtacag gaacgcacta tatctttcaa
     1621 agatgacggg aactacaaga cgcgtgctga agtcaagttt gaaggtgata cccttgttaa
     1681 tcgtatcgag ttaaaaggta ttgattttaa agaagatgga aacattctcg gacacaaact
     1741 cgagtacaac tataactcac acaatgtata catcacggca gacaaacaaa agaatggaat
     1801 caaagctaac ttcaaaattc gccacaacat tgaagatgga tccgttcaac tagcagacca
     1861 ttatcaacaa aatactccaa ttggcgatgg ccctgtcctt ttaccagaca accattacct
     1921 gtcgacacaa tctgcccttt cgaaagatcc caacgaaaag cgtgaccaca tggtccttct
     1981 tgagtttgta actgctgctg ggattacaca tggcatggat gagctctaca aataatgaat
     2041 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     2101 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     2161 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     2221 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     2281 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     2341 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     2401 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2461 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2521 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     2581 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     2641 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     2701 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     2761 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     2821 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     2881 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     2941 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3001 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     3061 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     3121 gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac
     3181 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact
     3241 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc
     3301 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg
     3361 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg
     3421 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt
     3481 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc
     3541 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag
     3601 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg
     3661 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc
     3721 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt
     3781 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg
     3841 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt
     3901 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct
     3961 tctgagcggg actctggggt tcgcgaaatg accgaccaag cgacgcccaa cctgccatca
     4021 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     4081 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     4141 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     4201 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     4261 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg
     4321 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     4381 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     4441 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     4501 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     4561 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     4621 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     4681 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     4741 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     4801 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     4861 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     4921 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     4981 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     5041 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     5101 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     5161 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     5221 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     5281 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     5341 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     5401 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     5461 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     5521 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     5581 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     5641 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     5701 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     5761 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     5821 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     5881 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     5941 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     6001 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     6061 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     6121 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     6181 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     6241 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     6301 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     6361 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     6421 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga
     6481 gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     6541 gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt
     6601 aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc
     6661 gttttgcgct gcttcgcgat gtacgggcca gatatacgcg tt
//
