LOCUS       Exported                8119 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Human TNF 3'UTR region cloned downstream of luciferase reporter 
            gene.
ACCESSION   .
VERSION     .
KEYWORDS    pmirGLO-3'UTR-TNF
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8119)
  AUTHORS   Emming S, Bianchi N, Polletti S, Balestrieri C, Leoni C, Montagner 
            S, Chirichella M, Delaleu N, Natoli G, Monticelli S
  TITLE     A molecular network regulating the proinflammatory phenotype of 
            human memory T lymphocytes.
  JOURNAL   Nat Immunol. 2020 Apr;21(4):388-399. doi: 10.1038/s41590-020-0622-8.
            Epub 2020 Mar 16.
  PUBMED    32205878
REFERENCE   2  (bases 1 to 8119)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8119
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     89..108
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      197..206
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             203..1855
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(2737..2755)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2792..2913
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2803..2822)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2857..2876
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3158..3515
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(3181..3201)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3366..3501
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3428..3447
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     regulatory      3560..3569
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             3566..4498
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     CDS             4550..5341
                     /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="IEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPV
                     LFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSS
                     HLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQG
                     LAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIAL
                     ATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4601..4620)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5211..5230
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5405..5453
                     /note="synthetic polyadenylation signal"
     promoter        5609..5713
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5714..6574
                     /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     complement(5932..5951)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6732..7320)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6812..6831)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    7436..7719
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     promoter        7776..156
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(7811..7834)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
ORIGIN
        1 cagaggctgg gaaggggtgg gtccgggggc gggctcaggg gcgggctcag gggcggggcg
       61 ggcgcccgaa ggtcctccgg aggcccggca ttctgcacgc ttcaaaagcg cacgtctgcc
      121 gcgctgttct cctcttcctc atctccgggc ctttcgacct gcagcccaag cttggcaatc
      181 cggtactgtt ggtaaagcca ccatggaaga tgccaaaaac attaagaagg gcccagcgcc
      241 attctaccca ctcgaagacg ggaccgccgg cgagcagctg cacaaagcca tgaagcgcta
      301 cgccctggtg cccggcacca tcgcctttac cgacgcacat atcgaggtgg acattaccta
      361 cgccgagtac ttcgagatga gcgttcggct ggcagaagct atgaagcgct atgggctgaa
      421 tacaaaccat cggatcgtgg tgtgcagcga gaatagcttg cagttcttca tgcccgtgtt
      481 gggtgccctg ttcatcggtg tggctgtggc cccagctaac gacatctaca acgagcgcga
      541 gctgctgaac agcatgggca tcagccagcc caccgtcgta ttcgtgagca agaaagggct
      601 gcaaaagatc ctcaacgtgc aaaagaagct accgatcata caaaagatca tcatcatgga
      661 tagcaagacc gactaccagg gcttccaaag catgtacacc ttcgtgactt cccatttgcc
      721 acccggcttc aacgagtacg acttcgtgcc cgagagcttc gaccgggaca aaaccatcgc
      781 cctgatcatg aacagtagtg gcagtaccgg attgcccaag ggcgtagccc taccgcaccg
      841 caccgcttgt gtccgattca gtcatgcccg cgaccccatc ttcggcaacc agatcatccc
      901 cgacaccgct atcctcagcg tggtgccatt tcaccacggc ttcggcatgt tcaccacgct
      961 gggctacttg atctgcggct ttcgggtcgt gctcatgtac cgcttcgagg aggagctatt
     1021 cttgcgcagc ttgcaagact ataagattca atctgccctg ctggtgccca cactatttag
     1081 cttcttcgct aagagcactc tcatcgacaa gtacgaccta agcaacttgc acgagatcgc
     1141 cagcggcggg gcgccgctca gcaaggaggt aggtgaggcc gtggccaaac gcttccacct
     1201 accaggcatc cgccagggct acggcctgac agaaacaacc agcgccattc tgatcacccc
     1261 cgaaggggac gacaagcctg gcgcagtagg caaggtggtg cccttcttcg aggctaaggt
     1321 ggtggacttg gacaccggta agacactggg tgtgaaccag cgcggcgagc tgtgcgtccg
     1381 tggccccatg atcatgagcg gctacgttaa caaccccgag gctacaaacg ctctcatcga
     1441 caaggacggc tggctgcaca gcggcgacat cgcctactgg gacgaggacg agcacttctt
     1501 catcgtggac cggctgaaga gcctgatcaa atacaagggc taccaggtag ccccagccga
     1561 actggagagc atcctgctgc aacaccccaa catcttcgac gccggggtcg ccggcctgcc
     1621 cgacgacgat gccggcgagc tgcccgccgc agtcgtcgtg ctggaacacg gtaaaaccat
     1681 gaccgagaag gagatcgtgg actatgtggc cagccaggtt acaaccgcca agaagctgcg
     1741 cggtggtgtt gtgttcgtgg acgaggtgcc taaaggactg accggcaagt tggacgcccg
     1801 caagatccgc gagattctca ttaaggccaa gaagggcggc aagatcgccg tgtaattcta
     1861 gttgtttaaa cgagctcgct agcggaggac gaacatccaa ccttcccaaa cgcctcccct
     1921 gccccaatcc ctttattacc ccctccttca gacaccctca acctcttctg gctcaaaaag
     1981 agaattgggg gcttagggtc ggaacccaag cttagaactt taagcaacaa gaccaccact
     2041 tcgaaacctg ggattcagga atgtgtggcc tgcacagtga agtgctggca accactaaga
     2101 attcaaactg gggcctccag aactcactgg ggcctacagc tttgatccct gacatctgga
     2161 atctggagac cagggagcct ttggttctgg ccagaatgct gcaggacttg agaagacctc
     2221 acctagaaat tgacacaagt ggaccttagg ccttcctctc tccagatgtt tccagacttc
     2281 cttgagacac ggagcccagc cctccccatg gagccagctc cctctattta tgtttgcact
     2341 tgtgattatt tattatttat ttattattta tttatttaca gatgaatgta tttatttggg
     2401 agaccggggt atcctggggg acccaatgta ggagctgcct tggctcagac atgttttccg
     2461 tgaaaacgga gctgaacaat aggctgttcc catgtagccc cctggcctct gtgccttctt
     2521 ttgattatgt tttttaaaat atttatctga ttaagttgtc taaacaatgc tgatttggtg
     2581 accaactgtc actcattgct gagcctctgc tccccagggg agttgtgtct gtaatcgccc
     2641 tactattcag tggcgagatc tagagtcgac ctgcaggcat gcaagctgat ccggctgcta
     2701 acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     2761 cccttggggc ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc
     2821 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     2881 tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg
     2941 tttcaggttc agggggagat gtgggaggtt tttttaagca agtaaaacct ctacaaatgt
     3001 ggtaaaatcg aattttaaca aaatattaac gcttacaatt tcctgatgcg gtattttctc
     3061 cttacgcatc tgtgcggtat ttcacaccgc atacgcggat ctgcgcagca ccatggcctg
     3121 aaataacctc tgaaagagga acttggttag gtaccttctg aggcggaaag aaccagctgt
     3181 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3241 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3301 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     3361 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3421 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     3481 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct tgattcttct gacacaacag
     3541 tctcgaacca aaggctggag ccaccatggc ttccaaggtg tacgaccccg agcaacgcaa
     3601 acgcatgatc actgggcctc agtggtgggc tcgctgcaag caaatgaacg tgctggactc
     3661 cttcatcaac tactatgatt ccgagaagca cgccgagaac gccgtgattt ttctgcatgg
     3721 taacgctgcc tccagctacc tgtggaggca cgtcgtgcct cacatcgagc ccgtggctag
     3781 atgcatcatc cctgatctga tcggaatggg taagtccggc aagagcggga atggctcata
     3841 tcgcctcctg gatcactaca agtacctcac cgcttggttc gagctgctga accttccaaa
     3901 gaaaatcatc tttgtgggcc acgactgggg ggcttgtctg gcctttcact actcctacga
     3961 gcaccaagac aagatcaagg ccatcgtcca tgctgagagt gtcgtggacg tgatcgagtc
     4021 ctgggacgag tggcctgaca tcgaggagga tatcgccctg atcaagagcg aagagggcga
     4081 gaaaatggtg cttgagaata acttcttcgt cgagaccatg ctcccaagca agatcatgcg
     4141 gaaactggag cctgaggagt tcgctgccta cctggagcca ttcaaggaga agggcgaggt
     4201 tagacggcct accctctcct ggcctcgcga gatccctctc gttaagggag gcaagcccga
     4261 cgtcgtccag attgtccgca actacaacgc ctaccttcgg gccagcgacg atctgcctaa
     4321 gatgttcatc gagtccgacc ctgggttctt ttccaacgct attgtcgagg gagctaagaa
     4381 gttccctaac accgagttcg tgaaggtgaa gggcctccac ttcagccagg aggacgctcc
     4441 agatgaaatg ggtaagtaca tcaagagctt cgtggagcgc gtgctgaaga acgagcagac
     4501 cggtggtggg agcggaggtg gcggatcagg tggcggaggc tccggaggga ttgaacaaga
     4561 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     4621 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     4681 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
     4741 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     4801 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     4861 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     4921 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     4981 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     5041 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt
     5101 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     5161 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     5221 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     5281 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     5341 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgatgg
     5401 ccgcaataaa atatctttat tttcattaca tctgtgtgtt ggttttttgt gtgaatcgat
     5461 agcgataagg atcctctttg cgcttgcgtt ttcccttgtc cagatagccc agtagctgac
     5521 attcatccgg ggtcagcacc gtttctgcgg actggctttc tacgtaatgg tttcttagac
     5581 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     5641 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     5701 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5761 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5821 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5881 gagttttcgc cccgaagaac gttttccaat gatgagcact ttcaaagttc tgctatgtgg
     5941 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     6001 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     6061 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     6121 tctgacaact atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     6181 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     6241 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     6301 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     6361 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     6421 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     6481 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6541 tgagataggt gcctcactga ttaagcattg gtaattcgaa atgaccgacc aagcgacgcc
     6601 caaccggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     6661 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     6721 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     6781 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     6841 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     6901 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     6961 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     7021 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     7081 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     7141 aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg
     7201 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     7261 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatttcaa
     7321 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     7381 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt atagtccgga
     7441 aatacaggaa cgcacgctgg atggcccttc gctgggatgg tgaaaccatg aaaaatggca
     7501 gcttcagtgg attaagtggg ggtaatgtgg cctgtaccct ctggttgcat aggtattcat
     7561 acggttaaaa tttatcaggc gcgattgcgg cagtttttcg ggtggtttgt tgccattttt
     7621 acctgtctgc tgccgtgatc gcgctgaacg cgttttagcg gtgcgtacaa ttaagggatt
     7681 atggtaaatc cacttactgt ctgccctcgt agccatcgag ataaaccgca gtactccggc
     7741 cacgatgcgt ccggcgtaga ggatcgagat ctaccgggta ggggaggcgc ttttcccaag
     7801 gcagtctgga gcatgcgctt tagcagcccc gctgggcact tggcgctaca caagtggcct
     7861 ctggcctcgc acacattcca catccaccgg taggcgccaa ccggctccgt tctttggtgg
     7921 ccccttcgcg ccaccttcta ctcctcccct agtcaggaag ttcccccccg ccccgcagct
     7981 cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc actagtctcg tgcagatgga
     8041 cagcaccgct gagcaatgga agcgggtagg cctttggggc agcggccaat agcagctttg
     8101 ctccttcgct ttctgggct
//
