LOCUS       pcDNA3-nuc-ExRai        6732 bp DNA     circular SYN 20-SEP-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6732)
  AUTHORS   Jin Zhang Lab
  TITLE     Direct Submission
  JOURNAL   Exported Sep 20, 2022 from SnapGene 6.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6732
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     CDS             922..939
                     /codon_start=1
                     /label=6His
                     /note="color: #008000"
                     /translation="HHHHHH"
     CDS             943..975
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /note="color: #cc99b2"
                     /translation="MASMTGGQQMG"
     CDS             979..1002
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /note="This feature has 2 segments:
                        1:  979 ..  987 / #cc99b2
                        2:  988 .. 1002 / #cc99b2 / enterokinase site"
                     /translation="DLYDDDDK"
     CDS             1009..1035
                     /codon_start=1
                     /label=AMPK substrate
                     /note="color: #999999"
                     /translation="MRRVATLVD"
     CDS             1072..1794
                     /codon_start=1
                     /label=cpGFP
                     /note="color: #008000"
                     /translation="NVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGPV
                     LLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEEL
                     FTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTY
                     GVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIEL
                     KGIDFKEDGNILGHKLEYN"
     CDS             1804..2226
                     /codon_start=1
                     /product="KFSQEQIGENIVCRVICTTGQIPIRDLSADISQVLKEKRSIKKVWTFGR
                     NPACDYHLGNISRLSNKHFQILLGEDGNLLLNDISTNGTWLNGQKVEKNSNQLLSQGDE
                     ITVGVGVESDILSLVIFINDKFKQCLEQNKVDR"
                     /label=FHA1
                     /note="color: #804002"
                     /translation="KFSQEQIGENIVCRVICTTGQIPIRDLSADISQVLKEKRSIKKVW
                     TFGRNPACDYHLGNISRLSNKHFQILLGEDGNLLLNDISTNGTWLNGQKVEKNSNQLLS
                     QGDEITVGVGVESDILSLVIFINDKFKQCLEQNKVDR"
     CDS             2236..2265
                     /codon_start=1
                     /label=NLS-Targeting
                     /note="color: #800040"
                     /translation="PKKKRKVEDA"
     promoter        complement(2284..2302)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     polyA_signal    2328..2552
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      2598..3026
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        3040..3370
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      3221..3356
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     CDS             3437..4231
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    4405..4526
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(4575..4591)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    4599..4615
                     /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)."
                     /note="color: #31849b"
     promoter        complement(4623..4653)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 4623 .. 4629 / #ffffff / -10
                        2: 4630 .. 4647 / #ffffff
                        3: 4648 .. 4653 / #ffffff / -35"
     protein_bind    4668..4689
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(4977..5565)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(5736..6596)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 5736 .. 6527 / #ccffcc
                        2: 6528 .. 6596 / #ccffcc / signal sequence
                     Cleavage site after base 6527"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6597..6701)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgg tgcggggttc tcatcatcat catcatcatg gtatggctag catgactggt
      961 ggacagcaaa tgggtcggga tctgtacgac gatgacgata aggatcccat gaggagagtg
     1021 gctaccctgg tggatctggg cggcaccggc ggcagcgagc tcagcttctg caacgtctat
     1081 atcaaggccg acaagcagaa gaacggcatc aaggcgaact tcaagatccg ccacaacatc
     1141 gaggacggcg gcgtgcagct cgcctaccac taccagcaga acacccccat cggcgacggc
     1201 cccgtgctgc tgcccgacaa ccactacctg agcgtgcagt ccaaactttc gaaagacccc
     1261 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     1321 ggcatggacg agctgtacaa gggcggtacc ggagggagca tggtgagcaa gggcgaggag
     1381 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1441 ttcagcgtgt ccggcgaggg tgagggcgat gccacctacg gcaagctgac cctgaagttc
     1501 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     1561 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1621 gccatgcccg aaggctacat ccaggagcgc accatcttct tcaaggacga cggcaactac
     1681 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1741 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caacttgctg
     1801 cataagtttt ctcaagaaca gatcggcgaa aacattgtgt gcagggtcat ttgtaccacg
     1861 ggtcaaattc ccatccgaga tttgtcagct gatatttcac aagtgcttaa ggaaaaacga
     1921 tccataaaga aagtttggac atttggtaga aacccagcct gtgactatca tttaggaaac
     1981 atttcaagac tgtcaaataa gcatttccaa atactactag gagaagacgg taacctttta
     2041 ttgaatgaca tttccactaa tgggacctgg ttaaatgggc aaaaagtcga gaagaacagc
     2101 aatcagttac tgtctcaagg tgatgaaata accgttggtg taggcgtgga atcagatatt
     2161 ttatctctgg tcattttcat aaacgacaaa tttaagcagt gcctcgagca gaacaaagtt
     2221 gatcgctatg aattccccaa aaagaagaga aaggtggaag acgcataatc tagagggccc
     2281 tattctatag tgtcacctaa atgctagagc tcgctgatca gcctcgactg tgccttctag
     2341 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     2401 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     2461 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     2521 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     2581 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     2641 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     2701 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc ggggcatccc
     2761 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     2821 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     2881 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     2941 ctattctttt gatttataag ggattttggg gatttcggcc tattggttaa aaaatgagct
     3001 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     3061 aagtccccag gctccccagg caggcagaag tatgcaaagc atgcatctca attagtcagc
     3121 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3181 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     3241 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     3301 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     3361 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg
     3421 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     3481 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     3541 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     3601 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     3661 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     3721 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     3781 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     3841 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     3901 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     3961 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4021 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4081 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4141 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4201 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4261 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4321 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4381 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     4441 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     4501 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     4561 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     4621 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     4681 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     4741 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     4801 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     4861 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     4921 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     4981 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5041 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5101 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5161 gcgctttctc aatgctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5221 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5281 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5341 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5401 tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5461 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5521 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     5581 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     5641 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     5701 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     5761 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     5821 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     5881 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     5941 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6001 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6061 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6121 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6181 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6241 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6301 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6361 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6421 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6481 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6541 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     6601 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     6661 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     6721 ccacctgacg tc
//
