LOCUS       Exported                7353 bp ds-DNA     circular SYN 17-JAN-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7353)
  AUTHORS   MIT
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Jan 21, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7353
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     109..128
                     /label=oSEQ054 pMXs-fwd-seq
     primer_bind     complement(198..211)
                     /label=oSEQ013 flipped_WPRE_rev
     protein_bind    222..246
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             285..296
                     /codon_start=1
                     /label=uORF
                     /note="/created_by=User"
                     /translation="TMG"
     CDS             297..308
                     /codon_start=1
                     /label=uORF
                     /note="/created_by=User"
                     /translation="TMG"
     CDS             309..320
                     /codon_start=1
                     /label=uORF
                     /note="/created_by=User"
                     /translation="TMG"
     primer_bind     321..359
                     /label=Lhx3_fwd
     primer_bind     321..354
                     /label=Lhx3_part1_fwd
     primer_bind     331..353
                     /label=Lhx3_1_fwd
     misc_feature    335..343
                     /label=Kozak sequence
                     /note="Partial (missing last G) vertebrate consensus 
                     sequence for strong initiation of translation (Kozak, 
                     1987)"
     primer_bind     338..359
                     /label=Lhx3_2_fwd
     primer_bind     339..359
                     /label=Lhx3_3_fwd
     misc_feature    341..1540
                     /label=mLhx3
                     /label=Lhx3
                     /note="Lhx3 isoform a from mouse (Mus musculus)"
     primer_bind     545..561
                     /label=oSEQ084 Lhx3mid_fwd
     primer_bind     complement(736..755)
                     /label=oSEQ104 Lhx3_rev
     primer_bind     complement(908..941)
                     /label=Lhx3_part1_rev
     primer_bind     922..949
                     /label=Lhx3_part2_fwd
     primer_bind     1448..1467
                     /label=oSEQ002 Lhx3_check_for
                     /note="25nm	STD"
     primer_bind     complement(1517..1540)
                     /label=Lhx3_part2_rev
     primer_bind     complement(1520..1544)
                     /label=Lhx3_1_rev
     primer_bind     complement(1520..1542)
                     /label=Lhx3_2_rev
     primer_bind     complement(1523..1540)
                     /label=Lhx3_rev
     primer_bind     complement(1523..1540)
                     /label=Lhx3-WPRE_rev
     primer_bind     complement(1523..1540)
                     /label=Lhx3_3_rev
     primer_bind     1540..1562
                     /label=oKL141
     primer_bind     1541..1563
                     /label=oKL166
     primer_bind     1541..1561
                     /label=P2A_fwd
     CDS             1550..1606
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     primer_bind     complement(1589..1616)
                     /label=Ngn2-P2A_rev_v2
     primer_bind     complement(1589..1606)
                     /label=P2A_rev
     primer_bind     complement(1591..1612)
                     /label=ggNgn2-P2A_rev
     primer_bind     1597..1625
                     /label=P2A-mRuby2_fwd
     primer_bind     1605..1625
                     /label=GG_mRuby2_fwd
     CDS             1607..2317
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     primer_bind     1607..1625
                     /label=mRuby2_fwd
                     /note="Fwd primer for mRuby2"
     primer_bind     1607..1625
                     /label=ggmRuby2_fwd
     primer_bind     1677..1696
                     /label=oSEQ062 mRuby2_fwd
     primer_bind     complement(1677..1696)
                     /label=oSEQ093 mRuby2_rev
     primer_bind     complement(2209..2228)
                     /label=oSEQ061 mRuby2_rev
     primer_bind     2243..2262
                     /label=OSEQ091 mRuby2_fwd_SEQ
     primer_bind     complement(2300..2324)
                     /label=ggmRuby2_rev
     primer_bind     complement(2302..2344)
                     /label=P2A-mRuby2_rev
     primer_bind     complement(2302..2322)
                     /label=mRuby2_rev
                     /note="Rev primer for mRuby2"
     primer_bind     complement(2302..2322)
                     /label=GG_mRuby2_rev
     primer_bind     complement(2302..2321)
                     /label=gemRuby2_rev
     primer_bind     complement(2325..2344)
                     /label=Lhx3_rev
     primer_bind     complement(2325..2344)
                     /label=Lhx3_part2_rev
     protein_bind    complement(2334..2358)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2375..2963
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2541..2560)
                     /label=oSEQ034 WPRE_rev
     primer_bind     2798..2818
                     /label=oSEQ048 phage-dest-fwd-2
     CDS             complement(2846..2857)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     2960..2977
                     /label=oSEQ013 flipped_WPRE_rev
     primer_bind     complement(3002..3020)
                     /label=oSEQ055 pMXs-rev-seq
     LTR             3143..3612
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     rep_origin      complement(3913..4501)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3993..4012)
                     /label=oSEQ067 pBR322-ori_fwd
     CDS             complement(4665..5525)
                     /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     5288..5307
                     /label=oSEQ066 Amp_rev
     primer_bind     complement(5501..5525)
                     /label=oSEQ083 Amp-fwd
     promoter        complement(5526..5630)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5572..5593
                     /label=oSEQ095 oKL157
     LTR             5660..6253
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    6316..6673
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             6732..7148
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(6926..6945)
                     /label=oSEQ082 pMX-gag-rev
     primer_bind     6926..6945
                     /label=oSEQ096 pMX-gag_fwd
     misc_feature    join(7158..7353,1..179)
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
ORIGIN
        1 tcggcgacac agtgtgggtc cgccgacacc agactaagaa cctagaacct cgctggaaag
       61 gaccttacac agtcctgctg accaccccca ccgccctcaa agtagacggc atcgcagctt
      121 ggatacacgc cgcccacgtg aaggctgccg accccggggg tggaccatcc tctagactgc
      181 cggatctagc tagttaatcg acggtatcga taagctatca aacaagtttg tacaaaaaag
      241 caggctccac catgggaacc aattcagtcg agtcgactgg atccaccatg ggttgaacca
      301 tgggttgaac catgggttga gatccggtac tactgccacc atgctgctag aagcagaact
      361 cgattgccac cgagagaggc ccggtgcccc tggagcttct gccctctgta ccttcagcag
      421 gactccagag atcccgatgt gtgcaggctg tgaccagcac atcttggacc gtttcatcct
      481 taaggctctg gaccgacatt ggcacagcaa gtgtctcaag tgcagtgact gccacgtccc
      541 tctggctgag cgctgcttca gccgcgggga gagcgtctac tgcaaagacg acttctttaa
      601 gcgcttcggg accaagtgcg ccgcatgcca gctgggcatc ccgcccacgc aggtggtgcg
      661 ccgcgcccag gacttcgtgt accacctgca ttgcttcgcc tgcgtggtct gcaagcggca
      721 gctggccacg ggcgacgagt tctacctcat ggaagacagc cggctggtgt gcaaggcgga
      781 ctacgaaaca gccaagcagc gagaagccga ggccacagcc aagcggccgc gcaccaccat
      841 caccgccaag cagctggaga cgctgaagag cgcctacaac acttcgccca agccggcgcg
      901 ccacgtgcgc gagcagctct cctccgagac tggcctggac atgcgagtgg tgcaggtgtg
      961 gttccagaat cgccgggcta aggaaaagag actgaagaaa gacgctggcc ggcagcgctg
     1021 gggacagtat ttccgcaata tgaagcgctc ccgcggcagt tccaagtccg acaaggacag
     1081 catccaggag ggacaagaca gcgacgccga agtctccttc actgatgagc cgtccatggc
     1141 tgacatgggg cctgctaatg gcctgtacag cagcctggga gagcctgccc ctgcgttggg
     1201 ccggcccgta ggaggcctgg gcagctttac cctggatcac ggaggcttga cgggtccaga
     1261 gcagtaccga gagctacgcc caggcagccc ctatggcatc cccccatctc ctgcagcccc
     1321 ccagagcctt cctggccccc agcctctcct ctccagcctg gtatacccag acaccaactt
     1381 gagccttgtc ccttcagggc ccccaggtgg acccccaccc atgagggtgc tggctggaaa
     1441 tgggcccagc tccgacctgt ccacagagag cagttctggc tacccagact ttcctgctag
     1501 ccctgcttcc tggctggatg aagtagacca tgctcagttc ggaagcggag ctactaactt
     1561 cagcctgctg aagcaggctg gagacgtgga ggagaaccct ggacctatgg tgtctaaggg
     1621 cgaagagctg atcaaggaaa atatgcgtat gaaggtggtc atggaaggtt cggtcaacgg
     1681 ccaccaattc aaatgcacag gtgaaggaga aggcaatccg tacatgggaa ctcaaaccat
     1741 gaggatcaaa gtcatcgagg gaggacccct gccatttgcc tttgacattc ttgccacgtc
     1801 gttcatgtat ggcagccgta cttttatcaa gtacccgaaa ggcattcctg atttctttaa
     1861 acagtccttt cctgagggtt ttacttggga aagagttacg agatacgaag atggtggagt
     1921 cgtcaccgtc atgcaggaca ccagccttga ggatggctgt ctcgtttacc acgtccaagt
     1981 cagaggggta aactttccct ccaatggtcc cgtgatgcag aagaagacca agggttggga
     2041 gcctaataca gagatgatgt atccagcaga tggtggtctg aggggataca ctcatatggc
     2101 actgaaagtt gatggtggtg gccatctgtc ttgctctttc gtaacaactt acaggtcaaa
     2161 aaagaccgtc gggaacatca agatgcccgg tatccatgcc gttgatcacc gcctggaaag
     2221 gttagaggaa agtgacaatg aaatgttcgt agtacaacgc gaacacgcag ttgccaagtt
     2281 cgccgggctt ggtggtggga tggacgagct gtacaagtaa cgctgatatc tagacccagc
     2341 tttcttgtac aaagtggttc gatagcttat cgataatcaa cctctggatt acaaaatttg
     2401 tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc
     2461 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta
     2521 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt
     2581 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca
     2641 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc
     2701 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     2761 gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg
     2821 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg
     2881 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat
     2941 ctccctttgg gccgcctccc cgcatcgata ccgtcgacga taaaataaaa gattttattt
     3001 agtctccaga aaaagggggg aatgaaagac cccacctgta ggtttggcaa gctagcttaa
     3061 gtaacgccat tttgcaaggc atggaaaaat acataactga gaatagagaa gttcagatca
     3121 aggtcaggaa cagatggaac agctgaatat gggccaaaca ggatatctgt ggtaagcagt
     3181 tcctgccccg gctcagggcc aagaacagat ggaacagctg aatatgggcc aaacaggata
     3241 tctgtggtaa gcagttcctg ccccggctca gggccaagaa cagatggtcc ccagatgcgg
     3301 tccagccctc agcagtttct agagaaccat cagatgtttc cagggtgccc caaggacctg
     3361 aaatgaccct gtgccttatt tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc
     3421 gcttctgctc cccgagctca ataaaagagc ccacaacccc tcactcgggg cgccagtcct
     3481 ccgattgact gagtcgcccg ggtacccgtg tatccaataa accctcttgc agttgcatcc
     3541 gacttgtggt ctcgctgttc cttgggaggg tctcctctga gtgattgact acccgtcagc
     3601 gggggtcttt cacatgcagc atgtatcaaa attaatttgg ttttttttct taagtattta
     3661 cattaaatgg ccatagttgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     3721 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     3781 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     3841 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     3901 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     3961 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4021 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     4081 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     4141 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     4201 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     4261 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     4321 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     4381 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     4441 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     4501 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     4561 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt tgcggccggc
     4621 cgcaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     4681 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     4741 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     4801 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     4861 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     4921 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     4981 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5041 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     5101 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     5161 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     5221 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     5281 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     5341 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     5401 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     5461 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     5521 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     5581 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     5641 cgaaaagtgc cacctgcata atgaaagacc ccacctgtag gtttggcaag ctagcttaag
     5701 taacgccatt ttgcaaggca tggaaaaata cataactgag aatagaaaag ttcagatcaa
     5761 ggtcaggaac agatggaaca gctgaatatg ggccaaacag gatatctgtg gtaagcagtt
     5821 cctgccccgg ctcagggcca agaacagatg gaacagctga atatgggcca aacaggatat
     5881 ctgtggtaag cagttcctgc cccggctcag ggccaagaac agatggtccc cagatgcggt
     5941 ccagccctca gcagtttcta gagaaccatc agatgtttcc agggtgcccc aaggacctga
     6001 aatgaccctg tgccttattt gaactaacca atcagttcgc ttctcgcttc tgttcgcgcg
     6061 cttctgctcc ccgagctcaa taaaagagcc cacaacccct cactcggcgc gccagtcctc
     6121 cgattgactg agtcgcccgg gtacccgtgt atccaataaa ccctcttgca gttgcatccg
     6181 acttgtggtc tcgctgttcc ttgggagggt ctcctctgag tgattgacta cccgtcagcg
     6241 ggggtctttc atttgggggc tcgtccggga tcgggagacc cctgcccagg gaccaccgac
     6301 ccaccaccgg gaggtaagct ggccagcaac ttatctgtgt ctgtccgatt gtctagtgtc
     6361 tatgactgat tttatgcgcc tgcgtcggta ctagttagct aactagctct gtatctggcg
     6421 gacccgtggt ggaactgacg agttcggaac acccggccgc aaccctggga gacgtcccag
     6481 ggacttcggg ggccgttttt gtggcccgac ctgagtccaa aaatcccgat cgttttggac
     6541 tctttggtgc acccccctaa taggagggat atgtggttct ggtaggagac gagaacctaa
     6601 aacagttccc gcctccgtct gaatttttgc tttcggtttg ggaccgaagc cgcgccgcgc
     6661 gtcttgtctg ctgcagcatc gttctgtgtt gtctctgtct gactgtgttt ctgtatttgt
     6721 ctgaaaatta gggccagact gttaccactc ccttaagttt gaccttaggt cactggaaag
     6781 atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa gaagagacgt tgggttacct
     6841 tctgctctgc agaatggcca acctttaacg tcggatggcc gcgagacggc acctttaacc
     6901 gagacctcat cacccaggtt aagatcaagg tcttttcacc tggcccgcat ggacacccag
     6961 accaggtccc ctacatcgtg acctgggaag ccttggcttt tgacccccct ccctgggtca
     7021 agccctttgt acaccctaag cctccgcctc ctcttcctcc atccgccccg tctctccccc
     7081 ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta tccagccctc actccttctc
     7141 taggcgcccc catatggcca tatgagatct tatatggggc acccccgccc cttgtaaact
     7201 tccctgaccc tgacatgaca agagttacta acagcccctc tctccaagct cacttacagg
     7261 ctctctactt agtccagcac gaagtctgga gacctctggc ggcagcctac caagaacaac
     7321 tggaccgacc ggtggtacct cacccttacc gag
//
