LOCUS       sequence-194085-        5238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of MCP-Tagged GPI anchor in Mammalian cells .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5238)
  AUTHORS   Ana Egana
  TITLE     Egana Lab NEB plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5238)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pMCP-GPI Control Plasmid
FEATURES             Location/Qualifiers
     source          1..5238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        81..272
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     229..249
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(435..1226,1227..1295))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1058..1077
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1296..1387)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(1406..1433)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     polyA_signal    1461..1582
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1472..1491)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1526..1545
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1856..1875
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1856..1874
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             complement(2022..2255)
                     /codon_start=1
                     /product="mutant acyl carrier protein (ACP) from E. coli"
                     /label=MCP
                     /note="The mutations (D36T and D39G) prevent recognition by
                     ACP synthase while allowing recognition by SFP synthase."
                     /translation="MSTIEERVKKIIGEQLGVKQEEVTNNASFVEDLGATSLGTVELVM
                     ALEEEFDTEIPDEEAEKITTVQAAIDYINGHQA"
     promoter        complement(2382..2400)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        complement(2446..2649)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2475..2495)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2650..2953
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3128..3716)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3208..3227)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4045..4092
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4117..4136
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4324..5118)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(4324..5118)
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4435..4454)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5045..5064
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(5153..5238,1..272))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(5167..5238,1..64)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 taaaaaaaat tagtcagcca tggggcggag aatgggcgga actgggcgga gttaggggcg
       61 ggatgggcgg agttaggggc gggactatgg ttgctgacta attgagatgc atgctttgca
      121 tacttctgcc tgctggggag cctggggact ttccacacct ggttgctgac taattgagat
      181 gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac cctaactgac
      241 acacattcca cagctggttc tttccgcctc aggattcttc ctttttcaat gtctgacgct
      301 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
      361 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
      421 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
      481 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
      541 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
      601 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
      661 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
      721 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
      781 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
      841 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
      901 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
      961 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     1021 cggcgaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc acatagcaga
     1081 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     1141 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     1201 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     1261 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     1321 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     1381 aaacaaaaga gtttgtagaa acgcaaaaag accatccgtc aggatggcct tctgcttatt
     1441 aacgcttaca atttacgcgt taagatacat tgatgagttt ggacaaacca caactagaat
     1501 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     1561 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     1621 gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg gtatggctga
     1681 ttatgatcat gaacagactg tgaggactga ggggcctgaa atgagccttg ggactgtgaa
     1741 tctaaaatac acaaacaatt agaatcagta gtttaacaca ttatacactt aaaaattgga
     1801 tctccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcgagtg aattgtaata
     1861 cgactcacta tagggcgaat tgggtacact tacctggtac cgggcccccc ctcgagtcag
     1921 gatccttagg gatgaaggct ccaggctgct gccagaaatg gagtcaccag cagaagaact
     1981 gttttctctg ataaggatgt cccaccattt tcaaggaatt ccgcctggtg gccgttgatg
     2041 taatcaatgg cagcctgaac ggtggtgatt ttctcagctt cttcgtccgg aatctcagta
     2101 tcaaactctt cttccagagc cattaccagc tcaacggtcc caagagaagt cgcgcccagg
     2161 tcttcaacga aagaagcatt gttggtaact tcttcctgct taacgcccag ctgttcgccg
     2221 ataattttct taacgcgttc ttcgatagtg ctcatagatc tgctatgacc tgaatggcag
     2281 aagacagcca ggacgagcag cagcccgaac aggacagacc ctccttggat tcccatggtg
     2341 atatcgcggc cgccaccgcg gtggagctcc agcttttgtt ccctttagtg agggttaatt
     2401 tcgagcttgg cgtaatcgct agcggatctg acggttcact aaaccagctc tgcttatata
     2461 gacctcccac cgtacacgcc taccgcccat ttgcgtcaat ggggcggagt tgttacgaca
     2521 ttttggaaag tcccgttgat tttggtgcca aaacaaactc ccattgacgt caatggggtg
     2581 gagacttgga aatccccgtg agtcaaaccg ctatccacgc ccattgatgt actgccaaaa
     2641 ccgcatcacc atggtaatag cgatgactaa tacgtagatg tactgccaag taggaaagtc
     2701 ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat tgacgtcaat
     2761 agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag tttaccgtaa
     2821 atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg gaacatacgt
     2881 cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc catttaccgt
     2941 aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg taattgatta
     3001 ctattaataa ctaatgcatg gcggtaatac ggttatccac agaatcaggg gataacgcag
     3061 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     3121 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     3181 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     3241 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     3301 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     3361 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     3421 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     3481 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     3541 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     3601 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     3661 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     3721 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     3781 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     3841 gttttaaatc aatctaaagt atatatgagt aacctgaggc tatggcaggg cctgccgccc
     3901 cgacgttggc tgcgagccct gggccttcac ccgaacttgg ggggtggggt ggggaaaagg
     3961 aagaaacgcg ggcgtattgg ccccaatggg gtctcggtgg ggtatcgaca gagtgccagc
     4021 cctgggaccg aaccccgcgt ttatgaacaa acgacccaac accgtgcgtt ttattctgtc
     4081 tttttattgc cgtcatagcg cgggttcctt ccggtattgt ctccttccgt gtttcagtta
     4141 gcctccccct agggtgggcg aagaactcca gcatgagatc cccgcgctgg aggatcatcc
     4201 agccggcgtc ccggaaaacg attccgaagc ccaacctttc atagaaggcg gcggtggaat
     4261 cgaaatctcg tgatggcagg ttgggcgtcg cttggtcggt catttcgaac cccagagtcc
     4321 cgctcagaag aactcgtcaa gaaggcgata gaaggcgatg cgctgcgaat cgggagcggc
     4381 gataccgtaa agcacgagga agcggtcagc ccattcgccg ccaagttctt cagcaatatc
     4441 acgggtagcc aacgctatgt cctgatagcg gtccgccaca cccagccggc cacagtcgat
     4501 gaatccagaa aagcggccat tttccaccat gatattcggc aagcaggcat cgccatgggt
     4561 cacgacgaga tcctcgccgt cgggcatgct cgccttgagc ctggcgaaca gttcggctgg
     4621 cgcgagcccc tgatgttctt cgtccagatc atcctgatcg acaagaccgg cttccatccg
     4681 agtacgtgct cgctcgatgc gatgtttcgc ttggtggtcg aatgggcagg tagccggatc
     4741 aagcgtatgc agccgccgca ttgcatcagc catgatggat actttctcgg caggagcaag
     4801 gtgagatgac aggagatcct gccccggcac ttcgcccaat agcagccagt cccttcccgc
     4861 ttcagtgaca acgtcgagca cagctgcgca aggaacgccc gtcgtggcca gccacgatag
     4921 ccgcgctgcc tcgtcttgca gttcattcag ggcaccggac aggtcggtct tgacaaaaag
     4981 aaccgggcgc ccctgcgctg acagccggaa cacggcggca tcagagcagc cgattgtctg
     5041 ttgtgcccag tcatagccga atagcctctc cacccaagcg gccggagaac ctgcgtgcaa
     5101 tccatcttgt tcaatcatgc gaaacgatcc tcatcctgtc tcttgatcga tctttgcaaa
     5161 agcctaggcc tccaaaaaag cctcctcact acttctggaa tagctcagag gccgaggcgg
     5221 cctcggcctc tgcataaa
//
