LOCUS       sequence-306228-        6585 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MSCV with IRES-Neomycin resistant gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6585)
  AUTHORS   Landry LG, Mann SE, Anderson AM, Nakayama M
  TITLE     Multiplex T-cell Stimulation Assay Utilizing a T-cell Activation 
            Reporter-based Detection System.
  JOURNAL   Bio Protoc. 2021 Jan 20;11(2):e3883. doi: 10.21769/BioProtoc.3883. 
            eCollection 2021 Jan 20.
  PUBMED    33732772
REFERENCE   2  (bases 1 to 6585)
  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: "Bio 
            Protoc."; date: "2021-01-20"; volume: "11(2):e3883. doi"; pages: " 
            10.21769/BioProtoc.3883. eCollection 2021 Jan 20"
COMMENT     Sequence Label: pMIII-Neo
FEATURES             Location/Qualifiers
     source          1..6585
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    64..405
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             472..888
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     818..840
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     856..872
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    957..1506
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1101..1118)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1328..1347
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             1516..2319
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1579..1598)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2189..2208
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     LTR             2463..2977
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    3139..3155
                     /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)."
     promoter        complement(join(3163..3169,3170..3187,3188..3193))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3208..3229
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3346..3363)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3517..4105)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3597..3616)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4276..5067,5068..5136))
                     /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     4899..4918
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5137..5241)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5309..5327
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5365..5387)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5487..5506
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5700..5722
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5850..5869
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             6069..6585
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tggaggttcc accgagattt ggagacccct gcccagggac caccgacccc cccgccggga
       61 ggtaagctgg ccagcggtcg tttcgtgtct gtctctgtct ttgtgcgtgt ttgtgccggc
      121 atctaatgtt tgcgcctgcg tctgtactag ttagctaact agctctgtat ctggcggacc
      181 cgtggtggaa ctgacgagtt ctgaacaccc ggccgcaacc ctgggagacg tcccagggac
      241 tttgggggcc gtttttgtgg cccgacctga ggaagggagt cgatgtggaa tccgaccccg
      301 tcaggatatg tggttctggt aggagacgag aacctaaaac agttcccgcc tccgtctgaa
      361 tttttgcttt cggtttggaa ccgaagccgc gcgtcttgtc tgctgcagcg ctgcagcatc
      421 gttctgtgtt gtctctgtct gactgtgttt ctgtatttgt ctgaaaatta gggccagact
      481 gttaccactc ccttaagttt gaccttaggt cactggaaag atgtcgagcg gatcgctcac
      541 aaccagtcgg tagatgtcaa gaagagacgt tgggttacct tctgctctgc agaatggcca
      601 acctttaacg tcggatggcc gcgagacggc acctttaacc gagacctcat cacccaggtt
      661 aagatcaagg tcttttcacc tggcccgcat ggacacccag accaggtccc ctacatcgtg
      721 acctgggaag ccttggcttt tgacccccct ccctgggtca agccctttgt acaccctaag
      781 cctccgcctc ctcttcctcc atccgccccg tctctccccc ttgaacctcc tcgttcgacc
      841 ccgcctcgat cctcccttta tccagccctc actccttctc taggcgccgg aattcagatc
      901 ttacgtagct agcggatccc aattgctcga gcgggatcaa ttccgccccc cccctaacgt
      961 tactggccga agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac
     1021 catattgccg tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag
     1081 cattcctagg ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa
     1141 ggaagcagtt cctctggaag cttcttgaag acaaacaacg tctgtagcga ccctttgcag
     1201 gcagcggaac cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga
     1261 tacacctgca aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag
     1321 agtcaaatgg ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc
     1381 ccattgtatg ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag
     1441 gttaaaaaaa cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg
     1501 ataataccat gaaccatggg atcggccatt gaacaagatg gattgcacgc aggttctccg
     1561 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     1621 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     1681 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     1741 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     1801 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     1861 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     1921 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     1981 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     2041 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     2101 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     2161 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     2221 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     2281 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag tcgacctgca ggcatgcaag
     2341 cttcaggtag ccggctaacg ttaacaaccg gtacctctag aactatagct agcatgcgca
     2401 aatttaaagc gctgatatcg ataaaataaa agattttatt tagtctccag aaaaaggggg
     2461 gaatgaaaga ccccacctgt aggtttggca agctagctta agtaacgcca ttttgcaagg
     2521 catggaaaat acataactga gaatagagaa gttcagatca aggttaggaa cagagagaca
     2581 gcagaatatg ggccaaacag gatatctgtg gtaagcagtt cctgccccgg ctcagggcca
     2641 agaacagatg gtccccagat gcggtcccgc cctcagcagt ttctagagaa ccatcagatg
     2701 tttccagggt gccccaagga cctgaaatga ccctgtgcct tatttgaact aaccaatcag
     2761 ttcgcttctc gcttctgttc gcgcgcttct gctccccgag ctcaataaaa gagcccacaa
     2821 cccctcactc ggcgcgccag tcctccgata gactgcgtcg cccgggtacc cgtgtatcca
     2881 ataaaccctc ttgcagttgc atccgacttg tggtctcgct gttccttggg agggtctcct
     2941 ctgagtgatt gactacccgt cagcgggggt ctttcatggg taacagtttc ttgaagttgg
     3001 agaacaacat tctgagggta ggagtcgaat attaagtaat cctgactcaa ttagccactg
     3061 ttttgaatcc acatactcca atactcctga aatagttcat tatggacagc gcagaaagag
     3121 ctggggagaa ttgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     3181 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3241 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     3301 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     3361 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     3421 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     3481 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     3541 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     3601 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     3661 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     3721 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     3781 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     3841 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     3901 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     3961 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     4021 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     4081 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     4141 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     4201 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     4261 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     4321 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     4381 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     4441 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     4501 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     4561 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     4621 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     4681 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     4741 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     4801 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     4861 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     4921 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     4981 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     5041 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     5101 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     5161 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     5221 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     5281 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     5341 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     5401 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     5461 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     5521 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
     5581 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     5641 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     5701 ccagtcacga cgttgtaaaa cgacggcgca aggaatggtg catgcaagga gatggcgccc
     5761 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     5821 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     5881 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggcg attagtccaa
     5941 tttgttaaag acaggatatc agtggtccag gctctagttt tgactcaaca atatcaccag
     6001 ctgaagccta tagagtacga gccatagata aaataaaaga ttttatttag tctccagaaa
     6061 aaggggggaa tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt
     6121 tgcaaggcat ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag
     6181 agagacagca gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc
     6241 agggccaaga acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca
     6301 tcagatgttt ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac
     6361 caatcagttc gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag
     6421 cccacaaccc ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt
     6481 attcccaata aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag
     6541 ggtctcctca gattgattga ctgcccacct cgggggtctt tcatt
//
