LOCUS       Exported                7397 bp DNA     circular SYN 12-MAY-2021
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 7397)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Mar 11, 2022 from SnapGene Viewer 6.0.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7397
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     enhancer        76..379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        380..583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     534..554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
                     /note="color: black; sequence: CGCAAATGGGCGGTAGGCGTG"
     mRNA            640..2460
                     /label=Calnexin
                     /note="color: #a6acb3; direction: RIGHT"
     primer_bind     complement(2450..2467)
                     /label=Vector.REV
                     /note="color: black; sequence: tcgactgcatggtggcga; added: 
                     2021-03-11"
     regulatory      2452..2460
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     primer_bind     2455..2473
                     /label=Fragment.FOR
                     /note="color: black; sequence: accatgcagtcgaccggca; added: 
                     2021-03-11"
     CDS             2476..3360
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /note="color: #cc99b2"
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     primer_bind     complement(3348..3368)
                     /label=Fragment.REV
                     /note="color: black; sequence: ttgctcacgccggaaatctcg; 
                     added: 2021-03-11"
     primer_bind     3352..3371
                     /label=Vector.FOR
                     /note="color: black; sequence: atttccggcgtgagcaaggg; added:
                     2021-03-11"
     CDS             3361..4077
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002)"
                     /label=mGold
                     /note="mammalian codon-optimized"
                     /note="This feature has 2 segments:
                        1: 3361 .. 3363 / #e1ff2f / 1a
                        2: 3364 .. 4077 / #e1ff2f"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTSLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     regulatory      3361
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     primer_bind     complement(3403..3424)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
                     /note="color: black; sequence: CGTCGCCGTCCAGCTCGACCAG"
     primer_bind     complement(3466..3484)
                     /label=EGFP-N-rev
                     /note="color: black; sequence: GCTTGCCGTAGGTGGCATC; added: 
                     2021-05-12"
     primer_bind     complement(3664..3683)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
                     /note="color: black; sequence: GTCTTGTAGTTGCCGTCGTC"
     primer_bind     4011..4032
                     /label=EGFP-C
                     /note="EGFP, forward primer"
                     /note="color: black; sequence: CATGGTCCTGCTGGAGTTCGTG"
     polyA_signal    4200..4321
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(4237..4256)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
                     /note="color: black; sequence: GAAATTTGTGATGCTATTGC"
     primer_bind     4291..4310
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
                     /note="color: black; sequence: GTGGTTTGTCCAAACTCATC"
     rep_origin      complement(4328..4783)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     complement(4465..4486)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
                     /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG"
     primer_bind     4677..4696
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
                     /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG"
     promoter        4810..4914
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     promoter        4916..5273
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     complement(4939..4959)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
                     /note="color: black; sequence: ACCCTAACTGACACACATTCC"
     rep_origin      5124..5259
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     primer_bind     5186..5205
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
                     /note="color: black; sequence: TATTTATGCAGAGGCCGAGG"
     primer_bind     complement(5209..5220)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
                     /note="color: black; sequence: TATTTATGCAGAGGCCGAGG"
     CDS             5308..6102
                     /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))"
                     /note="color: #ccffcc"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5362..5381)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
                     /note="color: black; sequence: GCCCAGTCATAGCCGAATAG"
     primer_bind     5972..5991
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
                     /note="color: black; sequence: CGTTGGCTACCCGTGATATT"
     primer_bind     complement(6290..6309)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
                     /note="color: black; sequence: TTGTCTCCTTCCGTGTTTCA"
     polyA_signal    6334..6381
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
                     /note="color: #a6acb3"
     rep_origin      6710..7298
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     7199..7218
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
                     /note="color: black; sequence: GGGAAACGCCTGGTATCTTT"
ORIGIN
        1 attaccgcca tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc
       61 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      121 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      181 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      241 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      301 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      361 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      421 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      481 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      541 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca
      601 gatccgctag cgctaccgga ctcagatctc gagaccgcca tggaagggaa gtggttgctg
      661 tgtatgttac tggtgcttgg aactgctatt gttgaggctc atgatggaca tgatgatgat
      721 gtgattgata ttgaggatga ccttgacgat gtcattgaag aggtagaaga ctcaaaacca
      781 gataccactg ctcctccttc atctcccaag gttacttaca aagctccagt tccaacaggg
      841 gaagtatatt ttgctgattc ttttgacaga ggaactctgt cagggtggat tttatccaaa
      901 gccaagaaag acgataccga tgatgaaatt gccaaatatg atggaaagtg ggaggtagag
      961 gaaatgaagg agtcaaagct tccaggtgat aaaggacttg tgttgatgtc tcgggccaag
     1021 catcatgcca tctctgctaa actgaacaag cccttcctgt ttgacaccaa gcctctcatt
     1081 gttcagtatg aggttaattt ccaaaatgga atagaatgtg gtggtgccta tgtgaaactg
     1141 ctttctaaaa caccagaact caacctggat cagttccatg acaagacccc ttatacgatt
     1201 atgtttggtc cagataaatg tggagaggac tataaactgc acttcatctt ccgacacaaa
     1261 aaccccaaaa cgggtatcta tgaagaaaaa catgctaaga ggccagatgc agatctgaag
     1321 acctatttta ctgataagaa aacacatctt tacacactaa tcttgaatcc agataatagt
     1381 tttgaaatac tggttgacca atctgtggtg aatagtggaa atctgctcaa tgacatgact
     1441 cctcctgtaa atccttcacg tgaaattgag gacccagaag accggaagcc cgaggattgg
     1501 gatgaaagac caaaaatccc agatccagaa gctgtcaagc cagatgactg ggatgaagat
     1561 gcccctgcta agattccaga tgaagaggcc acaaaacccg aaggctggtt agatgatgag
     1621 cctgagtacg tacctgatcc agacgcagag aaacctgagg attgggatga agacatggat
     1681 ggagaatggg aggctcctca gattgccaac cctagatgtg agtcagctcc tggatgtggt
     1741 gtctggcagc gacctgtgat tgacaacccc aattataaag gcaaatggaa gcctcctatg
     1801 attgacaatc ccagttacca gggaatctgg aaacccagga aaataccaaa tccagatttc
     1861 tttgaagatc tggaaccttt cagaatgact ccttttagtg ctattggttt ggagctgtgg
     1921 tccatgacct ctgacatttt ttttgacaac tttatcattt gtgctgatcg aagaatagtt
     1981 gatgattggg ccaatgatgg atggggcctg aagaaagctg ctgatggggc tgctgagcca
     2041 ggcgttgtgg ggcagatgaa cgaggcagct gaagagcgcc cgtggctgtg ggtagtctat
     2101 attctaactg tagcccttcc tgtgttcctg gttatcctct tctgctgttc tggaaagaaa
     2161 cagaccagtg gtatggagta taagaaaact gatgcacctc aaccggatgt gaaggaagag
     2221 gaagaagaga aggaagagga aaaggacaag ggagatgagg aggaggaagg agaagagaaa
     2281 cttgaagaga aacagaaaag tgatgctgaa gaagatggtg gcactgtcag tcaagaggag
     2341 gaagacagaa aacctaaagc agaggaggat gaaattttga acagatcacc aagaaacaga
     2401 aagccacgaa gagagggggg tgcgtcaggc ggcagcggat ctggaccggt cgccaccatg
     2461 cagtcgaccg gcagcgaaat cggtactggc tttccattcg acccccatta tgtggaagtc
     2521 ctgggcgagc gcatgcacta cgtcgatgtt ggtccgcgcg atggcacccc tgtgctgttc
     2581 ctgcacggta acccgacctc ctcctacgtg tggcgcaaca tcatcccgca tgttgcaccg
     2641 acccatcgct gcattgctcc agacctgatc ggtatgggca aatccgacaa accagacctg
     2701 ggttatttct tcgacgacca cgtccgcttc atggatgcct tcatcgaagc cctgggtctg
     2761 gaagaggtcg tcctggtcat tcacgactgg ggctccgctc tgggtttcca ctgggccaag
     2821 cgcaatccag agcgcgtcaa aggtattgca tttatggagt tcatccgccc tatcccgacc
     2881 tgggacgaat ggccagaatt tgcccgcgag accttccagg ccttccgcac caccgacgtc
     2941 ggccgcaagc tgatcatcga tcagaacgtt tttatcgagg gtacgctgcc gatgggtgtc
     3001 gtccgcccgc tgactgaagt cgagatggac cattaccgcg agccgttcct gaatcctgtt
     3061 gaccgcgagc cactgtggcg cttcccaaac gagctgccaa tcgccggtga gccagcgaac
     3121 atcgtcgcgc tggtcgaaga atacatggac tggctgcacc agtcccctgt cccgaagctg
     3181 ctgttctggg gcaccccagg cgttctgatc ccaccggccg aagccgctcg cctggccaaa
     3241 agcctgccta actgcaaggc tgtggacatc ggcccgggtc tgaatctgct gcaagaagac
     3301 aacccggacc tgatcggcag cgagatcgcg cgctggctgt ccacgctcga gatttccggc
     3361 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     3421 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     3481 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     3541 gtgaccagcc tgggctacgg cctgcagtgc ttcgcccgct accccgacca catgaagcag
     3601 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     3661 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     3721 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     3781 ctggagtaca actacaacag ccacaacgtc tatatcaccg ccgacaagca gaagaacggc
     3841 atcaaggcca acttcaagat ccgccacaac atcgaggacg gcggcgtgca gctcgccgac
     3901 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     3961 ctgagctacc agtccaagct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     4021 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaaagc
     4081 ggccgcgact ctagatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     4141 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     4201 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     4261 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     4321 aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     4381 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     4441 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     4501 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     4561 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     4621 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     4681 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     4741 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg
     4801 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     4861 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag
     4921 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     4981 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     5041 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     5101 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     5161 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     5221 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat
     5281 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     5341 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     5401 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     5461 gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc
     5521 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     5581 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     5641 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     5701 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     5761 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     5821 gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     5881 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     5941 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     6001 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     6061 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     6121 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     6181 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     6241 gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga
     6301 aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc
     6361 acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     6421 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     6481 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     6541 ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat ttttaattta
     6601 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     6661 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     6721 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     6781 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     6841 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     6901 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     6961 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     7021 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     7081 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     7141 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     7201 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     7261 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     7321 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     7381 attctgtgga taaccgt
//
