LOCUS       Exported                7361 bp ds-DNA     circular SYN 25-JAN-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 7361)
  AUTHORS   Nikhil Joshi Lab
  TITLE     Direct Submission
  JOURNAL   Exported Jul 17, 2024 from SnapGene 4.1.9
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7361
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     misc_feature    579..920
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             987..1403
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1333..1355
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1371..1387
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     1390..1430
                     /label=CMV FWD
     primer_bind     complement(1410..1430)
                     /label=GAG REV
     promoter        1466..1677
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     primer_bind     complement(1675..1699)
                     /label=Kozak REV
     primer_bind     1682..1717
                     /label=mKLF2 FWD
     regulatory      1691..1699
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     primer_bind     complement(1697..1743)
                     /label=KLF2_REV
     primer_bind     complement(1697..1717)
                     /label=MSCV_REV
     primer_bind     2145..2182
                     /label=KLF2_FWD
     primer_bind     2165..2182
                     /label=MSCV_FWD
     CDS             2765..2818
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     2823..2842
                     /label=mCherry FWD
     CDS             2825..3529
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWDASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     2825..2852
                     /label=MCHERRY FWD CTRL
     primer_bind     complement(2967..2985)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3235..3254
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(3566..3585)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     primer_bind     complement(3573..3615)
                     /label=mCherry REV
     primer_bind     3596..3617
                     /label=VECTOR FWD
     primer_bind     complement(3597..3619)
                     /label=MCHERRY REV
     LTR             3754..4268
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    4430..4446
                     /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(4454..4484)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4499..4520
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4637..4654)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4808..5396)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4888..4907)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5567..6427)
                     /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     6190..6209
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6428..6532)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6600..6618
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6656..6678)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6778..6797
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6991..7013
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7141..7160
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7360..7361,1..515)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc
      301 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
      361 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata
      421 aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag ggtctcctca
      481 gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga gatttggaga
      541 cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc ggtcgtttcg
      601 tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt
      661 actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac gagttctgaa
      721 cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt tgtggcccga
      781 cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt ctggtaggag
      841 acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tggaaccgaa
      901 gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc tgtctgactg
      961 tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta agtttgacct
     1021 taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat gtcaagaaga
     1081 gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga tggccgcgag
     1141 acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt tcacctggcc
     1201 cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg gcttttgacc
     1261 cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt cctccatccg
     1321 ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag
     1381 ccctcactcc ttctctaggc gccggaattg cttcgcgatg tacgggccag gaaaggagtg
     1441 ggaattggct ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag
     1501 ttggggggag gggtcggcaa ttgatccggt gcctagagaa ggtggcgcgg ggtaaactgg
     1561 gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata
     1621 agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacaggac
     1681 cggttctaga gccaccatgg cgctcagcga gcctatcttg cctagttttg ccacttttgc
     1741 atccccctgt gaacgaggat tgcaggaacg ctggcctcgc aacgaaccag aagctggagg
     1801 gaccgacgaa gacctcaaca atgtgttgga ttttatcctg agtatggggc tcgatggctt
     1861 gggggccgaa aacccaccag aaccacctcc acagccccct ccaccagcct tttattaccc
     1921 agaacccgga gcaccaccac catactcaat ccccgccgct tcacttggta cagaactcct
     1981 caggccagat ttggatcctc cacagggtcc cgcattgcac ggacgcttcc ttttggcccc
     2041 tccagggcga ctggtcaagg ccgagcctcc tgaagtggat ggtggaggct atgggtgcgc
     2101 tccagggctg gctcacgggc caagaggctt gaaacttgag ggcgctccag gtgcaaccgg
     2161 agcctgcatg cggggtcccg ccggccgccc cccgccgccc ccggacacgc cgccgctcag
     2221 ccccgacggc cccctgcgca tcccggcgtc cggtccacgc aacccgttcc cgccgccctt
     2281 cggtcccggc cccagcttcg gcggccccgg ccccgcgttg cactacgggc ctcccgcgcc
     2341 tggcgccttc ggtcttttcg aggacgcggc ggcagcggcg gcggcgctgg gcttggctcc
     2401 acctgccacg cgcggtctcc tcacgccgcc ctcgtccccg ctggagctgc tggaggccaa
     2461 gcccaaacgc ggccgccgct cctggccccg caagcgcgcc gccacacata cttgcagcta
     2521 caccaactgc ggcaagacct acaccaagag ctcgcaccta aaggcgcatc tgcgtacaca
     2581 cacaggtgag aagccttatc attgcaactg ggaaggatgc ggctggaagt tcgcgcgctc
     2641 tgacgagctt acccgccact accgaaagca cacaggtcac agaccctttc agtgccactt
     2701 gtgcgaccgg gccttctcca ggtccgacca cctggccttg cacatgaagc gacacatggc
     2761 tagcgagggc agaggaagtc ttctaacatg cggtgacgtg gaggagaatc ccggccctgg
     2821 taccgtgagc aagggcgagg aggataacat ggccatcatc aaggagttca tgcgcttcaa
     2881 ggtgcacatg gagggctccg tgaacggcca cgagttcgag atcgagggcg agggcgaggg
     2941 ccgcccctac gagggcaccc agaccgccaa gctgaaggtg accaagggtg gccccctgcc
     3001 cttcgcctgg gacatcctgt cccctcagtt catgtacggc tccaaggcct acgtgaagca
     3061 ccccgccgac atccccgact acttgaagct gtccttcccc gagggcttca agtgggagcg
     3121 cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc caggactcct ccctgcagga
     3181 cggcgagttc atctacaagg tgaagctgcg cggcaccaac ttcccctccg acggccccgt
     3241 aatgcagaag aagaccatgg gctgggatgc ctcctccgag cggatgtacc ccgaggacgg
     3301 cgccctgaag ggcgagatca agcagaggct gaagctgaag gacggcggcc actacgacgc
     3361 tgaggtcaag accacctaca aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt
     3421 caacatcaag ttggacatca cctcccacaa cgaggactac accatcgtgg aacagtacga
     3481 acgcgccgag ggccgccact ccaccggcgg catggacgag ctgtacaaga ccggttagta
     3541 atgagtttaa acgggggagg ctaactgaaa cacggaagga gacaataccg gaaggggccg
     3601 cgactctaga gtcgacctgc aggcatgcaa gcttcaggta gccggctaac gttaacaacc
     3661 ggtacctcta gaactatagc tagcatgcgc aaatttaaag cgctgatatc gataaaataa
     3721 aagattttat ttagtctcca gaaaaagggg ggaatgaaag accccacctg taggtttggc
     3781 aagctagctt aagtaacgcc attttgcaag gcatggaaaa tacataactg agaatagaga
     3841 agttcagatc aaggttagga acagagagac agcagaatat gggccaaaca ggatatctgt
     3901 ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtcccg
     3961 ccctcagcag tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg
     4021 accctgtgcc ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc
     4081 tgctccccga gctcaataaa agagcccaca acccctcact cggcgcgcca gtcctccgat
     4141 agactgcgtc gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt
     4201 gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg
     4261 tctttcatgg gtaacagttt cttgaagttg gagaacaaca ttctgagggt aggagtcgaa
     4321 tattaagtaa tcctgactca attagccact gttttgaatc cacatactcc aatactcctg
     4381 aaatagttca ttatggacag cgcagaaaga gctggggaga attgtgaaat tgttatccgc
     4441 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     4501 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     4561 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     4621 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     4681 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     4741 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     4801 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     4861 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     4921 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     4981 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     5041 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     5101 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     5161 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     5221 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     5281 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     5341 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5401 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     5461 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     5521 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     5581 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     5641 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     5701 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     5761 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     5821 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     5881 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     5941 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     6001 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     6061 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     6121 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     6181 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     6241 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     6301 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     6361 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     6421 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     6481 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     6541 cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa
     6601 ataggcgtat cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct
     6661 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     6721 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg
     6781 catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg
     6841 taaggagaaa ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag
     6901 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     6961 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcgc
     7021 aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac
     7081 catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc
     7141 ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac
     7201 gatgcgtccg gcgtagaggc gattagtcca atttgttaaa gacaggatat cagtggtcca
     7261 ggctctagtt ttgactcaac aatatcacca gctgaagcct atagagtacg agccatagat
     7321 aaaataaaag attttattta gtctccagaa aaagggggga a
//
