LOCUS       pNP-843_pCDNA3.1       10120 bp DNA     circular SYN 09-SEP-2022
DEFINITION  New Assembly 10120bp ds-DNA circular SYN  9-SEP-2022.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10120)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 9, 2022 from SnapGene Viewer 6.1.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..10120
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    72..296
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      342..770
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        784..1113
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      964..1099
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1180..1974
                     /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"
     polyA_signal    2148..2269
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2318..2334)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2342..2358
                     /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(2366..2396)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2411..2432
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(2720..3305)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3476..4336)
                     /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"
     promoter        complement(4337..4441)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4707..5086
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5087..5290
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        5335..5353
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    5384..5389
                     /label=Kozak
     misc_feature    5387..5443
                     /label=A3 SP
     CDS             5447..5485
                     /codon_start=1
                     /product="rationally designed epitope tag that forms a 
                     stable alpha-helix (Gotzke et al., 2019)"
                     /label=ALFA-tag
                     /note="It may be beneficial to flank the ALFA-tag sequence 
                     with proline residues to preserve the helical structure."
                     /translation="SRLEEELRRRLTE"
     misc_feature    5489..5545
                     /label=lk-HiBiT-lk
     misc_feature    5546..10117
                     /label=ADGRL3
ORIGIN
        1 ctgcagatat ccagcacagt ggcggccgct cgagtctaga gggcccgttt aaacccgctg
       61 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
      121 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
      181 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
      241 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
      301 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
      361 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
      421 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
      481 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
      541 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
      601 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
      661 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
      721 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
      781 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
      841 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
      901 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
      961 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     1021 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     1081 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     1141 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     1201 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     1261 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     1321 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     1381 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     1441 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     1501 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     1561 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     1621 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     1681 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     1741 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     1801 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     1861 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     1921 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     1981 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     2041 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     2101 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     2161 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     2221 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     2281 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     2341 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     2401 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     2461 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     2521 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     2581 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     2641 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     2701 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     2761 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     2821 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     2881 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     2941 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     3001 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     3061 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     3121 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     3181 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3241 ccaccgctgg tagcggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     3301 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     3361 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     3421 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     3481 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     3541 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     3601 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     3661 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     3721 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     3781 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     3841 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     3901 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     3961 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     4021 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     4081 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     4141 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     4201 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     4261 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     4321 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     4381 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     4441 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct
     4501 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc
     4561 tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca
     4621 aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg
     4681 cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc
     4741 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     4801 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     4861 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     4921 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     4981 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     5041 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     5101 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     5161 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     5221 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     5281 aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac
     5341 gactcactat agggagaccc aagctggcta gcgtttaaac ttagccatgt ggccaccaca
     5401 gctactcatc ctcacgatgc ttttagcacc tgtagttcat ggtccaagca gactggagga
     5461 ggagctgaga agaagactga ccgagccagg cagcagcggc gtgagcggct ggcggctgtt
     5521 caagaagatt agcggcagca gcggcggcaa gcacaatgag agacatccag ccctcgctgc
     5581 tccactgcga cacgctgagc gcagcccagg aggcgctctc cctcccagac atctccttca
     5641 gcagccagcg gcagagcgct caacggctca tcgaggacag gggccacgcg gagctgccag
     5701 aggagtccgt ggaccaggtg ccccaggagc acagattgca gcccaagctt tcagccgcgc
     5761 cccaattccc atggcagtgg tccgcagaga gctctcctgt gagagctacc ccattgagct
     5821 ccgttgtccg ggcacagacg tcatcatgat cgagagcgcc aactacggga ggaccgatga
     5881 caagatctgt gactcggacc ctgctcagat ggagaacatt cggtgttatc tgccagatgc
     5941 ctataagatt atgtctcaaa gatgcaataa cagaacccag tgtgcagtgg tggcaggtcc
     6001 tgatgtattt ccagacccat gtccgggaac atataaatac cttgaagtgc agtatgaatg
     6061 tgtcccttat aaagtggaac aaaaagtttt tctttgtcct ggactgctaa aaggagtgta
     6121 ccagagtgaa cacttgtttg aatctgacca ccaatctggg gcatggtgca aggaccctct
     6181 acaggcttct gacaagattt actatatgcc ctggactccc tacagaactg acaccctgac
     6241 agagtattcg tccaaagatg acttcattgc tggaaggcca acaactacct acaagctccc
     6301 tcacagagtg gatggcactg gatttgtagt atacgatggt gccctcttct tcaacaagga
     6361 gcggacaagg aacattgtaa agtttgattt gaggactagg ataaagagtg gagaggcaat
     6421 catagcaaat gctaattacc atgacacctc cccataccga tggggtggca agtccgacat
     6481 agacttggca gtggatgaaa atggattatg ggtaatctat gcaacagaac agaacaatgg
     6541 caagattgtc attagccagt tgaaccctta caccctacgg attgagggga catgggacac
     6601 tgcctatgat aaaaggtcag cttccaatgc attcatgatt tgtgggattc tgtatgtggt
     6661 caaatctgta tatgaggatg atgacaacga ggccaccggt aataagattg actacattta
     6721 caacactgac caaagcaagg atagcttggt ggatgtaccc tttcccaact cttaccagta
     6781 catagcagct gtggattaca atcccaggga caatctgctc tatgtgtgga ataactacca
     6841 cgttgtgaaa tactctttgg acttcgggcc tctggatagt agatcagggc cggtacatca
     6901 tggacaagtc tcctacatct ctccaccaat tcacctcgac tctgaactag aaaggccccc
     6961 tgtcagaggg atttctacca caggatccct gggtatggga agcacgacca ccagcaccac
     7021 cctccggacc acaacctgga acataggcag gagtaccacc gcatccttgc cgggcagaag
     7081 aaaccgcagt accagcacgc catcccccgc ggtagaggtg ctggatgacg tcaccacaca
     7141 cctgccctcg gcagcctccc aaatcccagc tatggaagag agctgcgagg ctgtggaagc
     7201 ccgagaaatc atgtggttta agaccagaca ggggcaggta gcaaagcagc catgcccagc
     7261 aggaaccata ggtgtatcaa cttacctatg tcttgctcct gatggaatat gggaccccca
     7321 aggaccagat ctcagcaact gctcttctcc ttgggtcaat cacataacac agaagctgaa
     7381 atccggagaa acagctgcca atattgccag agagctagca gaacagacaa gaaatcactt
     7441 gaacgctggg gacatcacct actctgtccg tgccatggac cagctggttg gcctcttgga
     7501 tgtccagctc aggaatttga caccaggggg aaaagacagc gcagcccgca gcttgaacaa
     7561 gcttcagaaa agagagcgct cttgcagagc ctatgtccag gcgatggtgg agacagttaa
     7621 caacctcctt cagccacaag ctttgaatgc ctggagagac ctgacgacaa gtgatcaact
     7681 gcgtgcggcc accatgttgc ttgacactgt ggaggagagc gctttcgtgt tagctgataa
     7741 ccttttgaag accgacattg tcagggagaa tacagacaat attcagttag aagttgcaag
     7801 actgagcacg gaaggaaacc tagaagatct aaaattccca gagaacatgg gccacggaag
     7861 taccatccag ctctcagcaa acacattaaa gcaaaatggc cggaatggag agattagagt
     7921 ggcttttgtc ctgtataata acctgggtcc ttatttgtct acggagaatg ccagtatgaa
     7981 gttgggcaca gaagctatgt ccacaaatca ctcggttatc gtcaattccc ctgttattac
     8041 agcagcaata aataaggaat tcagtaataa agtgtatttg gctgatcctg tggtatttac
     8101 tgttaaacac atcaagcagt cagaagaaaa tttcaaccct aactgttcat tttggagcta
     8161 ttccaagcgc acaatgacag gttattggtc aacacaaggc tgtcgactcc tgacaaccaa
     8221 caagacacat actacatgct cctgtaacca cctcaccaac tttgcagtat tgatggcaca
     8281 tgtggaagtt aagcacagcg atgcagtcca cgatcttctt ctggatgtga tcacgtgggt
     8341 tggcatcttg ctgtcccttg tctgtctcct gatttgcatc ttcacattct gcttcttccg
     8401 gggactccag agtgaccgca acaccattca caagaacctc tgcatcagcc tgttcgtggc
     8461 agaactgctc ttcctgattg gaatcaacag aaccgaccaa ccaattgcct gtgcagtgtt
     8521 cgcggctctt ttgcatttct tcttcttggc ggccttcacc tggatgtttc tagaaggggt
     8581 gcagctgtat atcatgctgg tggaggtctt tgagagcgag cattcccgta ggaagtactt
     8641 ttatctggtc ggctatggga tgcccgcgct catcgtggcc gtttctgcgg ccgtcgacta
     8701 caggagctac ggaacagaca aagtatgttg gcttcgcctt gacacctact tcatttggag
     8761 ttttatagga ccggcgacct tgataattat gctgaatgtc atcttccttg ggattgcttt
     8821 atacaaaatg tttcaccata ctgccatact gaaacctgaa tcaggctgtc ttgacaatat
     8881 caactatgag gataacagac ccttcatcaa gtcatgggtt ataggtgcaa tagctctgct
     8941 ctgcctgtta ggattgacct gggcctttgg actcatgtat attaatgaaa gcacagtcat
     9001 catggcgtat ctcttcacca ttttcaattc tctacaggga atgtttatat tcattttcca
     9061 ctgtgttcta cagaagaagg tacggaaaga gtatgggaaa tgcctccgca cgcattgctg
     9121 tagtgggaaa agcacggaga gttcgattgg ctcagggaaa acatctggtt ctcgaactcc
     9181 aggacggtac tccacaggct cgcagagccg gattcggaga atgtggaatg acactgtccg
     9241 aaagcagtca gagtcgtcct tcatcaccgg agacataaac agctcagcat cgctcaacag
     9301 agagccctac agagagacaa aggggcttct gaacaatgcc agggatacaa gtgtcatgga
     9361 tactctacca ctgaatggta accatggcaa cagttacagc attgctggcg gcgaatacct
     9421 gagcaactgt gtgcaaatta tagaccgtgg ctataaccac aacgagaccg ccctagagaa
     9481 aaagatccta aaggaactca cttccaacta tatcccttca tacctgaaca accacgagcg
     9541 ctccagcgaa cagaaccgga atatgatgaa caaactggtg aacaacttag gcagcgggag
     9601 tgaagatgac gccatcgtcc tggatgacgc cgcgtccttt aaccacgagg agagtctggg
     9661 cctggaactc attcacgagg aatccgatgc tcccttgctg cccccgaggg tttactccac
     9721 ggataaccac cagccacacc attacagcag gaggcggttc ccccaggacc acagcgagag
     9781 cttcttccct ctgctaaccg acgagcacac agaagacctg cagtcaccgc acagggactc
     9841 tctgtacacc agcatgccag ccctggccgg cgtgcccgct gcagacagtg tgaccaccag
     9901 cacccagacc gaagccgcag cggccaaggg tggtgacgcc gaagatgttt actacaaaag
     9961 catgccaaac ctgggctcca gaaaccacgt gcacccgctg cacgcctact accagctggg
    10021 gcgcggcagc agcgacggat tcatagttcc tcctaacaaa gatggggcct ctccggaggg
    10081 gacttccaaa ggacccgcgc acttggtcac tagtctataa
//
