LOCUS       sequence-390122-        8783 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express mini-Agrin in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8783)
  TITLE     Jeff Miner Lab Plasmids
REFERENCE   2  (bases 1 to 8783)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pCI_E01_mini-mAgrin-Myc-His-WPRE__LG3
FEATURES             Location/Qualifiers
     source          1..8783
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    40..323
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     enhancer        514..893
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        894..1097
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1048..1068
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          1233..1365
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1348..1367
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1409..1428
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1409..1427
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1528..1537
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3934..3954
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             3979..4008
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4018..4047
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4057..4086
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4096..4125
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4135..4164
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4192..4209
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    4246..4834
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4299..4319)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4717..4728)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4922..4940)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    4977..5098
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4988..5007)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5042..5061
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5343..5700
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5343..5534
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5366..5386)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5551..5686
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5731..6768
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             6747..6755
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    6792..6840
                     /note="synthetic polyadenylation signal"
     promoter        6996..7100
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7101..7169,7170..7961)
                     /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(7319..7338)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(8119..8707)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8199..8218)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gattatcaaa aaggatcttc acctagatcc ttttatagtc cggaaataca ggaacgcacg
       61 ctggatggcc cttcgctggg atggtgaaac catgaaaaat ggcagcttca gtggattaag
      121 tgggggtaat gtggcctgta ccctctggtt gcataggtat tcatacggtt aaaatttatc
      181 aggcgcgatt gcggcagttt ttcgggtggt ttgttgccat ttttacctgt ctgctgccgt
      241 gatcgcgctg aacgcgtttt agcggtgcgt acaattaagg gattatggta aatccactta
      301 ctgtctgccc tcgtagccat cgagataaac cgcagtactc cggccacgat gcgtccggcg
      361 tagaggatcg agatcttcaa tattggccat tagccatatt attcattggt tatatagcat
      421 aaatcaatat tggctattgg ccattgcata cgttgtatct atatcataat atgtacattt
      481 atattggctc atgtccaata tgaccgccat gttggcattg attattgact agttattaat
      541 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
      601 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      661 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      721 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtccgcccc
      781 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttac
      841 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
      901 ggttttggca gtacaccaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
      961 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     1021 aatgtcgtaa taaccccgcc ccgttgacgc aaatgggcgg taggcgtgta cggtgggagg
     1081 tctatataag cagagctggt ttagtgaacc gtcagatcac tagaagcttt attgcggtag
     1141 tttatcacag ttaaattgct aacgcagtca gtgcttctga cacaacagtc tcgaacttaa
     1201 gctgcagaag ttggtcgtga ggcactgggc aggtaagtat caaggttaca agacaggttt
     1261 aaggagacca atagaaactg ggcttgtcga gacagagaag actcttgcgt ttctgatagg
     1321 cacctattgg tcttactgac atccactttg cctttctctc cacaggtgtc cactcccagt
     1381 tcaattacag ctcttaaggc tagagtatta atacgactca ctatagggct agcaaagcga
     1441 tcgccgaact cacctatctc cccaacacct aataacattc aatcactctt tccactaacc
     1501 acctatctac atcaccaccc accagccgcc accatggtcc gcccgcggct gagcttcccg
     1561 gccccgctgc tgccgctgct gctgctcctg gcggcggccg cgccagccgt gccgggcgcc
     1621 agcgggacgt gcccggagcg cgccttggag cggcgcgagg aggaggcgaa cgtggtgctc
     1681 accggcacag tggaggagat cctcaacgtg gacccggtgc agcacaccta ctcctgcaag
     1741 gttcgagtgt ggcggtactt gaaaggcaaa gatgtggtgg cccaagagag cctgctggat
     1801 ggtggcaaca aggtggtaat tggtggcttt ggagaccccc tcatctgcga caaccaggtg
     1861 tccaccgggg acactagaat cttctttgtg aacccagcac caccctacct gtggccagcc
     1921 cacaagaatg agttgatgct caactccagc cttatgcgga tcactttgcg gaacctggaa
     1981 gaggtggagt tctgtgtaga agacaaacct gggatccact tcacagcagc gccttctatg
     2041 cctcctgatg tgtgcagggg aatgttatgt ggctttggtg ctgtgtgtga acctagtgtt
     2101 gaggatccag gccgggcctc ctgtgtgtgc aagaagaatg tctgccctgc tatggtagct
     2161 cctgtgtgtg gctcagatgc ttccacctat agcaacgagt gtgagctaca gcgtgcacag
     2221 tgcaaccagc aacggcgcat ccgcctactc cgccaagggc catgtgggtc ccgggacccc
     2281 tgtgccaatg tgaccgctag cactccaggc ccgcaacggc ccccaaagtc ctgtgattcc
     2341 cagccttgcc tccacggagg tacctgccag gacctggatt ctggcaaggg tttcagctgc
     2401 agctgtactg caggcagggc tggcactgtc tgtgagaaag tgcaactccc ctctgtgcca
     2461 gcttttaagg gccactcctt cttggccttc cccaccctcc gagcctacca cacgctgcgt
     2521 ctggcactag aattccgggc gctggagaca gagggactgc tgctctacaa tggcaatgca
     2581 cgtggcaaag atttcctggc tctggctctg ttggatggtc atgtacagtt caggttcgac
     2641 acgggctcag ggccggcggt gctaacaagc ttagtgccag tggaaccggg acggtggcac
     2701 cgcctcgagt tgtcacggca ttggcggcag ggcacacttt ctgtggatgg cgaggctcct
     2761 gttgtaggtg aaagtcccag tgggactgat ggcctcaact tggacacgaa gctctatgtg
     2821 ggtggtctcc cagaagaaca agttgccacg gtgcttgatc ggacctctgt gggcatcggc
     2881 ctgaaaggat gcattcgtat gttggacatc aacaaccagc agctggagct gagcgattgg
     2941 cagagggctg tggttcaaag ctctggtgtg ggggaatgcg gagaccatcc ctgctcacct
     3001 aacccctgcc atggcggggc cctctgccag gccctggagg ctggcgtgtt cctctgtcag
     3061 tgtccacctg gccgctttgg cccaacttgt gcagatgaaa agaacccctg ccaaccgaac
     3121 ccctgccacg ggtcagcccc ctgccatgtg ctttccaggg gtggggccaa gtgtgtgtgc
     3181 cccctgggac gcagtggttc cttctgtgag acagtcctgg agaatgctgg ctcccggccc
     3241 ttcctggctg actttaatgg cttctcctac ctggaactga aaggcttgca caccttcgag
     3301 agagacctag gggagaagat ggcgctggag atggtgttct tggctcgtgg gcccagtggc
     3361 ttactcctct acaatgggca gaagacggat ggcaaggggg actttgtatc cctggccctg
     3421 cataaccggc acctagagtt ccgctatgac cttggcaagg gggctgcaat catcaggagc
     3481 aaagagccca tagccctggg cacctgggtt agggtattcc tggaacgaaa tggccgcaag
     3541 ggtgcccttc aagtgggtga tgggccccgt gtgctagggg aatctccggt cccgcacacc
     3601 atgctcaacc tcaaggagcc cctctatgtg gggggagctc ctgacttcag caagctggct
     3661 cggggcgctg cagtggcctc cggctttgat ggtgccatcc agctggtgtc tctaagaggc
     3721 catcagctgc tgactcagga gcatgtgttg cgggcagtag atgtagcgcc ttttgcaggc
     3781 cacccttgta cccaggccgt ggacaacccc tgccttaatg ggggctcctg tatcccgagg
     3841 gaagccactt atgagtgcct gtgtcctggg ggcttctctg ggctgcactg cgagaagggg
     3901 atagttgaga agtcagtggg ggaccccact ctcgaggatc tgtactttca gagcatcgat
     3961 ggaagtggta aagctatgga gcaaaagctc atttctgaag aggacttgaa tgaaatggag
     4021 caaaagctca tttctgaaga ggacttgaat gaaatggagc aaaagctcat ttctgaagag
     4081 gacttgaatg aaatggagca aaagctcatt tctgaagagg acttgaatga aatggagcaa
     4141 aagctcattt ctgaagagga cttgaatgaa atggagagct tgggcgacct ccatcatcac
     4201 catcaccact aggggatatc agctgaattc tagaaattca tcgacaatca acctctggat
     4261 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     4321 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     4381 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     4441 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     4501 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     4561 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     4621 tccgtggtgt tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg tgttgccacc
     4681 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     4741 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     4801 acgagtcgga tctccctttg ggccgcctcc ccgcccaggt tccttgggcc gcgactctag
     4861 ggcatgcaag ctgatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
     4921 accgctgagc aataactagc ataacccctt ggggcggccg cttcgagcag acatgataag
     4981 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     5041 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     5101 caacaacaat tgcattcatt ttatgtttca ggttcagggg gagatgtggg aggttttttt
     5161 aagcaagtaa aacctctaca aatgtggtaa aatcgaattt taacaaaata ttaacgctta
     5221 caatttcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     5281 cggatctgcg cagcaccatg gcctgaaata acctctgaaa gaggaacttg gttaggtacc
     5341 ttctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     5401 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     5461 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     5521 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     5581 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
     5641 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     5701 aagctcgatt cttctgacac tagcgccacc atgaagaagc ccgaactcac cgctaccagc
     5761 gttgaaaaat ttctcatcga gaagttcgac agtgtgagcg acctgatgca gttgtcggag
     5821 ggcgaagaga gccgagcctt cagcttcgat gtcggcggac gcggctatgt actgcgggtg
     5881 aatagctgcg ctgatggctt ctacaaagac cgctacgtgt accgccactt cgccagcgct
     5941 gcactaccca tccccgaagt gttggacatc ggcgagttca gcgagagcct gacatactgc
     6001 atcagtagac gcgcccaagg cgttactctc caagacctcc ccgaaacaga gctgcctgct
     6061 gtgttacagc ctgtcgccga agctatggat gctattgccg ccgccgacct cagtcaaacc
     6121 agcggcttcg gcccattcgg gccccaaggc atcggccagt acacaacctg gcgggatttc
     6181 atttgcgcca ttgctgatcc ccatgtctac cactggcaga ccgtgatgga cgacaccgtg
     6241 tccgccagcg tagctcaagc cctggacgaa ctgatgctgt gggccgaaga ctgtcccgag
     6301 gtgcgccacc tcgtccatgc cgacttcggc agcaacaacg tcctgaccga caacggccgc
     6361 atcaccgccg taatcgactg gtccgaagct atgttcgggg acagtcagta cgaggtggcc
     6421 aacatcttct tctggcggcc ctggctggct tgcatggagc agcagactcg ctacttcgag
     6481 cgccggcatc ccgagctggc cggcagccct cgtctgcgag cctacatgct gcgcatcggc
     6541 ctggatcagc tctaccagag cctcgtggac ggcaacttcg acgatgctgc ctgggctcaa
     6601 ggccgctgcg atgccatcgt ccgcagcggg gccggcaccg tcggtcgcac acaaatcgct
     6661 cgccggagcg cagccgtatg gaccgacggc tgcgtcgagg tgctggccga cagcggcaac
     6721 cgccggccca gtacacgacc gcgcgctaag gaggtaggtc gagtttagac tctagccatc
     6781 acgatggccg caataaaata tctttatttt cattacatct gtgtgttggt tttttgtgtg
     6841 aatcgatagc gataaggatc ctctttgcgc ttgcgttttc ccttgtccag atagcccagt
     6901 agctgacatt catccggggt cagcaccgtt tctgcggact ggctttctac gtaatggttt
     6961 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     7021 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     7081 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     7141 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     7201 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     7261 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttc aaagttctgc
     7321 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     7381 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     7441 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     7501 acttacttct gacaactatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     7561 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     7621 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     7681 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     7741 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     7801 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     7861 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     7921 agatcgctga gataggtgcc tcactgatta agcattggta attcgaaatg accgaccaag
     7981 cgacgcccaa ccggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     8041 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     8101 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     8161 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     8221 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     8281 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     8341 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     8401 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     8461 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     8521 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     8581 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     8641 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     8701 atttcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     8761 acgttaaggg attttggtca tga
//
