LOCUS       sequence-390123-        7307 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 7307)
  TITLE     Jeff Miner Lab Plasmids
REFERENCE   2  (bases 1 to 7307)
  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__LG1-3
FEATURES             Location/Qualifiers
     source          1..7307
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    34..317
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     enhancer        508..887
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        888..1091
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1042..1062
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          1227..1359
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1342..1361
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1403..1422
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1403..1421
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1522..1531
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2452..2472
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             2497..2526
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2536..2565
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2575..2604
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2614..2643
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2653..2682
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2710..2727
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    2764..3352
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2817..2837)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3235..3246)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3440..3458)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3495..3616
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3506..3525)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3560..3579
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3861..4218
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3861..4052
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3884..3904)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4069..4204
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4249..5286
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             5265..5273
                     /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    5310..5358
                     /note="synthetic polyadenylation signal"
     promoter        5514..5618
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5619..5687,5688..6479)
                     /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(5837..5856)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6637..7225)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6717..6736)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 caaaaaggat cttcacctag atccttttat agtccggaaa tacaggaacg cacgctggat
       61 ggcccttcgc tgggatggtg aaaccatgaa aaatggcagc ttcagtggat taagtggggg
      121 taatgtggcc tgtaccctct ggttgcatag gtattcatac ggttaaaatt tatcaggcgc
      181 gattgcggca gtttttcggg tggtttgttg ccatttttac ctgtctgctg ccgtgatcgc
      241 gctgaacgcg ttttagcggt gcgtacaatt aagggattat ggtaaatcca cttactgtct
      301 gccctcgtag ccatcgagat aaaccgcagt actccggcca cgatgcgtcc ggcgtagagg
      361 atcgagatct tcaatattgg ccattagcca tattattcat tggttatata gcataaatca
      421 atattggcta ttggccattg catacgttgt atctatatca taatatgtac atttatattg
      481 gctcatgtcc aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat
      541 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
      601 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      661 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      721 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg
      781 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact
      841 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
      901 ggcagtacac caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
      961 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     1021 gtaataaccc cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     1081 taagcagagc tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc
     1141 acagttaaat tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca
     1201 gaagttggtc gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag
     1261 accaatagaa actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta
     1321 ttggtcttac tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt
     1381 acagctctta aggctagagt attaatacga ctcactatag ggctagcaaa gcgatcgccg
     1441 aactcaccta tctccccaac acctaataac attcaatcac tctttccact aaccacctat
     1501 ctacatcacc acccaccagc cgccaccatg gtccgcccgc ggctgagctt cccggccccg
     1561 ctgctgccgc tgctgctgct cctggcggcg gccgcgccag ccgtgccggg cgccagcggg
     1621 acgtgcccgg agcgcgcctt ggagcggcgc gaggaggagg cgaacgtggt gctcaccggc
     1681 acagtggagg agatcctcaa cgtggacccg gtgcagcaca cctactcctg caaggttcga
     1741 gtgtggcggt acttgaaagg caaagatgtg gtggcccaag agagcctgct ggatggtggc
     1801 aacaaggtgg taattggtgg ctttggagac cccctcatct gcgacaacca ggtgtccacc
     1861 ggggacacta gaatcttctt tgtgaaccca gcaccaccct acctgtggcc agcccacaag
     1921 aatgagttga tgctcaactc cagccttatg cggatcactt tgcggaacct ggaagaggtg
     1981 gagttctgtg tagaagacaa acctgggatc cacttcacag cagcgccttc tatgcctcct
     2041 gatgtgtgca ggggaatgtt atgtggcttt ggtgctgtgt gtgaacctag tgttgaggat
     2101 ccaggccggg cctcctgtgt gtgcaagaag aatgtctgcc ctgctatggt agctcctgtg
     2161 tgtggctcag atgcttccac ctatagcaac gagtgtgagc tacagcgtgc acagtgcaac
     2221 cagcaacggc gcatccgcct actccgccaa gggccatgtg ggtcccggga cccctgtgcc
     2281 aatgtgaccg ctagcactcc aggcccgcaa cggcccccaa agtcctgtga ttcccagcct
     2341 tgcctccacg gaggtacctg ccaggacctg gattctggca agggtttcag ctgcagctgt
     2401 actgcaggca gggctggcac tgtctgtgag aaagtgcagc tccccactct cgaggatctg
     2461 tactttcaga gcatcgatgg aagtggtaaa gctatggagc aaaagctcat ttctgaagag
     2521 gacttgaatg aaatggagca aaagctcatt tctgaagagg acttgaatga aatggagcaa
     2581 aagctcattt ctgaagagga cttgaatgaa atggagcaaa agctcatttc tgaagaggac
     2641 ttgaatgaaa tggagcaaaa gctcatttct gaagaggact tgaatgaaat ggagagcttg
     2701 ggcgacctcc atcatcacca tcaccactag gggatatcag ctgaattcta gaaattcatc
     2761 gacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt
     2821 gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc
     2881 cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag
     2941 ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc
     3001 actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc
     3061 cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg
     3121 ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt tccatggctg
     3181 ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc
     3241 ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt
     3301 cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc gcccaggttc
     3361 cttgggccgc gactctaggg catgcaagct gatccggctg ctaacaaagc ccgaaaggaa
     3421 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcggccgct
     3481 tcgagcagac atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg
     3541 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     3601 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     3661 gatgtgggag gtttttttaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgaatttta
     3721 acaaaatatt aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg
     3781 tatttcacac cgcatacgcg gatctgcgca gcaccatggc ctgaaataac ctctgaaaga
     3841 ggaacttggt taggtacctt ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     3901 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3961 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4021 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     4081 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     4141 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     4201 ggcctaggct tttgcaaaaa gctcgattct tctgacacta gcgccaccat gaagaagccc
     4261 gaactcaccg ctaccagcgt tgaaaaattt ctcatcgaga agttcgacag tgtgagcgac
     4321 ctgatgcagt tgtcggaggg cgaagagagc cgagccttca gcttcgatgt cggcggacgc
     4381 ggctatgtac tgcgggtgaa tagctgcgct gatggcttct acaaagaccg ctacgtgtac
     4441 cgccacttcg ccagcgctgc actacccatc cccgaagtgt tggacatcgg cgagttcagc
     4501 gagagcctga catactgcat cagtagacgc gcccaaggcg ttactctcca agacctcccc
     4561 gaaacagagc tgcctgctgt gttacagcct gtcgccgaag ctatggatgc tattgccgcc
     4621 gccgacctca gtcaaaccag cggcttcggc ccattcgggc cccaaggcat cggccagtac
     4681 acaacctggc gggatttcat ttgcgccatt gctgatcccc atgtctacca ctggcagacc
     4741 gtgatggacg acaccgtgtc cgccagcgta gctcaagccc tggacgaact gatgctgtgg
     4801 gccgaagact gtcccgaggt gcgccacctc gtccatgccg acttcggcag caacaacgtc
     4861 ctgaccgaca acggccgcat caccgccgta atcgactggt ccgaagctat gttcggggac
     4921 agtcagtacg aggtggccaa catcttcttc tggcggccct ggctggcttg catggagcag
     4981 cagactcgct acttcgagcg ccggcatccc gagctggccg gcagccctcg tctgcgagcc
     5041 tacatgctgc gcatcggcct ggatcagctc taccagagcc tcgtggacgg caacttcgac
     5101 gatgctgcct gggctcaagg ccgctgcgat gccatcgtcc gcagcggggc cggcaccgtc
     5161 ggtcgcacac aaatcgctcg ccggagcgca gccgtatgga ccgacggctg cgtcgaggtg
     5221 ctggccgaca gcggcaaccg ccggcccagt acacgaccgc gcgctaagga ggtaggtcga
     5281 gtttagactc tagccatcac gatggccgca ataaaatatc tttattttca ttacatctgt
     5341 gtgttggttt tttgtgtgaa tcgatagcga taaggatcct ctttgcgctt gcgttttccc
     5401 ttgtccagat agcccagtag ctgacattca tccggggtca gcaccgtttc tgcggactgg
     5461 ctttctacgt aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     5521 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     5581 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     5641 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     5701 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     5761 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     5821 gcactttcaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     5881 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     5941 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     6001 gtgataacac tgcggccaac ttacttctga caactatcgg aggaccgaag gagctaaccg
     6061 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     6121 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     6181 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     6241 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     6301 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     6361 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     6421 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaat
     6481 tcgaaatgac cgaccaagcg acgcccaacc ggtatcagct cactcaaagg cggtaatacg
     6541 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     6601 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     6661 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     6721 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     6781 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     6841 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     6901 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     6961 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     7021 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac
     7081 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     7141 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     7201 tacgcgcaga aaaaaaggat ttcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     7261 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattat
//
