LOCUS       Exported                7152 bp ds-DNA     circular SYN 26-OCT-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7152)
  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..7152
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          2968
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    45..386
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             453..869
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     799..821
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     837..853
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     846..868
                     /label=EcoRI Insert F
     misc_feature    847..875
                     /label=Gibson EcoRI site
     misc_feature    872..1271
                     /label=sgRNA casette frag
     promoter        883..1123
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     883..903
                     /label=hU6 forward psv
     primer_bind     1113..1134
                     /label=U6 Forward
     misc_feature    1148..1266
                     /label=sgRNA scaffold
     misc_feature    1214..1235
                     /label=10X Capture Sequence 1
     primer_bind     complement(1267..1289)
                     /label=sgRNA cassette reverse
     misc_feature    1295..1324
                     /label=Gibson Arm 1
     primer_bind     complement(1298..1321)
                     /label=Primer 1
     promoter        1325..1824
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     1325..1346
                     /label=Primer 5
     primer_bind     complement(1360..1383)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1753..1771
                     /label=PS_Vector_3
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     1757..1776
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     misc_feature    1825..1854
                     /label=Gibson Arm 5
     CDS             1864..2916
                     /codon_start=1
                     /label=hCD2
                     /translation="MSFPCKFVASFLLIFNVSSKGAVSKEITNALETWGALGQDINLDI
                     PSFQMSDDIDDIKWEKTSDKKKIAQFRKEKETFKEKDTYKLFKNGTLKIKHLKTDDQDI
                     YKVSIYDTKGKNVLEKIFDLKIQERVSKPKISWTCIKTTLTCEVMNGTDPELNLYQDGK
                     HLKLSQRVITHKWTTSLSAKFKCTAGNKVSKESSVEPVSCPEKGLDIYLIIGICGGGSL
                     LMVFVALLVFYITKRKKQRSRRNDEELETRAHRVATEERGRKPHQIPASTPQNPATSQH
                     PPPPPGHRSQAPSHRPPPPGHRVQHQPQKRPPAPSGTQVHQQKGPPLPRPRVQPKPPHG
                     AAENSLSPSSN"
     primer_bind     2159..2181
                     /label=PSV-hCD2
     primer_bind     2270..2292
                     /label=PS_Vector_4
     primer_bind     complement(2542..2568)
                     /label=Primer 6
     primer_bind     2773..2793
                     /label=PS_Vector_5
     primer_bind     2940..2969
                     /label=PSV P2 V2
     LTR             3011..3525
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    3687..3703
                     /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(3711..3741)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3756..3777
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3894..3911)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4065..4653)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4145..4164)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4824..5684)
                     /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     5447..5466
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5685..5789)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5857..5875
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5913..5935)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6035..6054
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6248..6270
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6398..6417
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             6617..7152
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tggagacccc tgcccaggga ccaccgaccc ccccgccggg aggtaagctg gccagcggtc
       61 gtttcgtgtc tgtctctgtc tttgtgcgtg tttgtgccgg catctaatgt ttgcgcctgc
      121 gtctgtacta gttagctaac tagctctgta tctggcggac ccgtggtgga actgacgagt
      181 tctgaacacc cggccgcaac cctgggagac gtcccaggga ctttgggggc cgtttttgtg
      241 gcccgacctg aggaagggag tcgatgtgga atccgacccc gtcaggatat gtggttctgg
      301 taggagacga gaacctaaaa cagttcccgc ctccgtctga atttttgctt tcggtttgga
      361 accgaagccg cgcgtcttgt ctgctgcagc gctgcagcat cgttctgtgt tgtctctgtc
      421 tgactgtgtt tctgtatttg tctgaaaatt agggccagac tgttaccact cccttaagtt
      481 tgaccttagg tcactggaaa gatgtcgagc ggatcgctca caaccagtcg gtagatgtca
      541 agaagagacg ttgggttacc ttctgctctg cagaatggcc aacctttaac gtcggatggc
      601 cgcgagacgg cacctttaac cgagacctca tcacccaggt taagatcaag gtcttttcac
      661 ctggcccgca tggacaccca gaccaggtcc cctacatcgt gacctgggaa gccttggctt
      721 ttgacccccc tccctgggtc aagccctttg tacaccctaa gcctccgcct cctcttcctc
      781 catccgcccc gtctctcccc cttgaacctc ctcgttcgac cccgcctcga tcctcccttt
      841 atccagccct cactccttct ctaggcgccg gaattcaacg tagagggcct atttcccatg
      901 attccttcat atttgcatat acgatacaag gctgttagag agataattag aattaatttg
      961 actgtaaaca caaagatatt agtacaaaat acgtgacgta gaaagtaata atttcttggg
     1021 tagtttgcag ttttaaaatt atgttttaaa atggactatc atatgcttac cgtaacttga
     1081 aagtatttcg atttcttggc tttatatatc ttgtggaaag gacgaaacac cgggtcttcg
     1141 agaagacgtt taagagctaa gctggaaaca gcatagcaag tttaaataag gctagtccgt
     1201 tatcaacttg gccgctttaa ggccggtcct agcaaggcca agtggcaccg agtcggtgct
     1261 ttttttggat ccatctctgt ctgactgtgt ttctgatcag tgtgagggag tgtaaagctg
     1321 gtttgggtag gggaggcgct tttcccaagg cagtctggag catgcgcttt agcagccccg
     1381 ctgggcactt ggcgctacac aagtggcctc tggcctcgca cacattccac atccaccggt
     1441 aggcgccaac cggctccgtt ctttggtggc cccttcgcgc caccttctac tcctccccta
     1501 gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca ggacgtgaca aatggaagta
     1561 gcacgtctca ctagtctcgt gcagatggac agcaccgctg agcaatggaa gcgggtaggc
     1621 ctttggggca gcggccaata gcagctttgc tccttcgctt tctgggctca gaggctggga
     1681 aggggtgggt ccgggggcgg gctcaggggc gggctcaggg gcggggcggg cgcccgaagg
     1741 tcctccggag gcccggcatt ctgcacgctt caaaagcgca cgtctgccgc gctgttctcc
     1801 tcttcctcat ctccgggcct ttcgaaacgt tgttgtttgg ggttgaatta ctctctcgag
     1861 accatgagct ttccatgtaa atttgtagcc agcttccttc tgattttcaa tgtttcttcc
     1921 aaaggtgcag tctccaaaga gattacgaat gccttggaaa cctggggtgc cttgggtcag
     1981 gacatcaact tggacattcc tagttttcaa atgagtgatg atattgacga tataaaatgg
     2041 gaaaaaactt cagacaagaa aaagattgca caattcagaa aagagaaaga gactttcaag
     2101 gaaaaagata catataagct atttaaaaat ggaactctga aaattaagca tctgaaaacc
     2161 gatgatcagg atatctacaa ggtatcaata tatgatacaa aaggaaaaaa tgtgttggaa
     2221 aaaatatttg atttgaagat tcaagagagg gtctcaaaac caaagatctc ctggacttgt
     2281 atcaaaacaa ccctgacctg tgaggtaatg aatggaactg accccgaatt aaacctgtat
     2341 caagatggga aacatctaaa actttctcag agggtcatca cacacaagtg gaccaccagc
     2401 ctgagtgcaa aattcaagtg cacagcaggg aacaaagtca gcaaggaatc cagtgtcgag
     2461 cctgtcagct gtccagagaa aggtctggac atctatctca tcattggcat atgtggagga
     2521 ggcagcctct tgatggtctt tgtggcactg ctcgttttct atatcaccaa aaggaaaaaa
     2581 cagaggagtc ggagaaatga tgaggagctg gagacaagag cccacagagt agctactgaa
     2641 gaaaggggcc ggaagcccca ccaaattcca gcttcaaccc ctcagaatcc agcaacttcc
     2701 caacatcctc ctccaccacc tggtcatcgt tcccaggcac ctagtcatcg tcccccgcct
     2761 cctggacacc gtgttcagca ccagcctcag aagaggcctc ctgctccgtc gggcacacaa
     2821 gttcaccagc agaaaggccc gcccctcccc agaccccgag ttcagccaaa acctccccat
     2881 ggggcagcag aaaactcatt gtccccttcc tctaattagg cttgcgcggc cgctaggtac
     2941 ctttaagacc aatgacttac aaggatcgat aaaataaaag attttattta gtctccagaa
     3001 aaagggggga atgaaagacc ccacctgtag gtttggcaag ctagcttaag taacgccatt
     3061 ttgcaaggca tggaaaatac ataactgaga atagagaagt tcagatcaag gttaggaaca
     3121 gagagacagc agaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct
     3181 cagggccaag aacagatggt ccccagatgc ggtcccgccc tcagcagttt ctagagaacc
     3241 atcagatgtt tccagggtgc cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa
     3301 ccaatcagtt cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga
     3361 gcccacaacc cctcactcgg cgcgccagtc ctccgataga ctgcgtcgcc cgggtacccg
     3421 tgtatccaat aaaccctctt gcagttgcat ccgacttgtg gtctcgctgt tccttgggag
     3481 ggtctcctct gagtgattga ctacccgtca gcgggggtct ttcatgggta acagtttctt
     3541 gaagttggag aacaacattc tgagggtagg agtcgaatat taagtaatcc tgactcaatt
     3601 agccactgtt ttgaatccac atactccaat actcctgaaa tagttcatta tggacagcgc
     3661 agaaagagct ggggagaatt gtgaaattgt tatccgctca caattccaca caacatacga
     3721 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3781 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3841 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3901 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     3961 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4021 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4081 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4141 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4201 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4261 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4321 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4381 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4441 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4501 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4561 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4621 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4681 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4741 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4801 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4861 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4921 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     4981 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5041 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5101 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5161 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5221 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5281 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5341 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5401 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5461 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5521 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5581 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5641 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5701 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5761 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     5821 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     5881 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     5941 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     6001 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
     6061 gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     6121 cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     6181 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
     6241 gggttttccc agtcacgacg ttgtaaaacg acggcgcaag gaatggtgca tgcaaggaga
     6301 tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg aaacaagcgc
     6361 tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc
     6421 cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggcgat
     6481 tagtccaatt tgttaaagac aggatatcag tggtccaggc tctagttttg actcaacaat
     6541 atcaccagct gaagcctata gagtacgagc catagataaa ataaaagatt ttatttagtc
     6601 tccagaaaaa ggggggaatg aaagacccca cctgtaggtt tggcaagcta gcttaagtaa
     6661 cgccattttg caaggcatgg aaaatacata actgagaata gagaagttca gatcaaggtt
     6721 aggaacagag agacagcaga atatgggcca aacaggatat ctgtggtaag cagttcctgc
     6781 cccggctcag ggccaagaac agatggtccc cagatgcggt cccgccctca gcagtttcta
     6841 gagaaccatc agatgtttcc agggtgcccc aaggacctga aatgaccctg tgccttattt
     6901 gaactaacca atcagttcgc ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa
     6961 taaaagagcc cacaacccct cactcggcgc gccagtcctc cgatagactg cgtcgcccgg
     7021 gtacccgtat tcccaataaa gcctcttgct gtttgcatcc gaatcgtgga ctcgctgatc
     7081 cttgggaggg tctcctcaga ttgattgact gcccacctcg ggggtctttc atttggaggt
     7141 tccaccgaga tt
//
