LOCUS       DD-pGEMHE-Mm_CEN        5323 bp DNA     circular SYN 19-DEC-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5323)
  AUTHORS   121 Lynch Laboratories
  TITLE     Direct Submission
  JOURNAL   Exported Mar 24, 2025 from SnapGene 7.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5323
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     promoter        complement(63..167)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     551..570
                     /label=QCH_seq_primer_1
                     /note="color: black; sequence: CCTCTTCGCTATTACGCCAG"
     primer_bind     641..657
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     promoter        664..682
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     5'UTR           714..756
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
                     /note="color: #a6acb3; direction: RIGHT"
     primer_bind     764..796
                     /label=InFu_Mm_CENP-T_F
                     /note="color: black; sequence: 
                     attccccggtgctagATGGCGGACCTCAGCTTT"
     misc_feature    779..2323
                     /label=Mm_CENP-T
                     /note="color: #00ffff"
     primer_bind     779..796
                     /label=MmCT_F
                     /note="color: black; sequence: ATGGCGGACCTCAGCTTT; added: 
                     2022-12-19"
     misc_feature    991..1009
                     /label=siRNA_1 target
                     /note="color: #a6acb3"
     misc_feature    1040..1058
                     /label=siRNA_2 target
                     /note="color: #a6acb3"
     misc_feature    1092..1110
                     /label=siRNA_3 target
                     /note="color: #a6acb3"
     primer_bind     1169..1190
                     /label=Mm_CENP-T_seq_1
                     /note="color: black; sequence: CTGGAGCTACACCTTCCTGAAC"
     primer_bind     1751..1772
                     /label=Mm_CENP-T_seq_2
                     /note="color: black; sequence: GCAGAACCGAGAGAAGCAGTAG"
     misc_feature    2015..2287
                     /label=HFD
                     /note="color: #cc99ff"
     primer_bind     complement(2057..2087)
                     /label=A1294G antisense
                     /note="color: black; sequence: 
                     ctatctcaagagccgccttctccacgggcat"
     primer_bind     2057..2087
                     /label=A1294G
                     /note="color: black; sequence: 
                     atgcccgtggagaaggcggctcttgagatag"
     primer_bind     2145..2177
                     /label=T1388C
                     /note="color: black; sequence: 
                     cttctcatgctggccgtaagactgtgaagccag"
     primer_bind     complement(2145..2177)
                     /label=T1388C antisense
                     /note="color: black; sequence: 
                     ctggcttcacagtcttacggccagcatgagaag"
     primer_bind     2160..2201
                     /label=T1408G_T1409A
                     /note="color: black; sequence: 
                     gtaagattgtgaagccagaggacttagagctgctgatgcgac"
     primer_bind     complement(2160..2201)
                     /label=T1408G_T1409A antisense
                     /note="color: black; sequence: 
                     gtcgcatcagcagctctaagtcctctggcttcacaatcttac"
     primer_bind     complement(2301..2338)
                     /label=InFu_Mm_CENP-T_R
                     /note="color: black; sequence: 
                     tccgccaccgACCGGCTGTGCAGGGAAGACAGAATTGC"
     primer_bind     complement(2301..2323)
                     /label=MmCT_R
                     /note="color: black; sequence: CTGTGCAGGGAAGACAGAATTGC; 
                     added: 2022-12-19"
     primer_bind     2324..2343
                     /label=Fragment.FOR3
                     /note="color: black; sequence: 
                     tgaggagaGAACCGGTcggtggcggaggtgg"
     misc_feature    2330..2344
                     /label=5Gly
                     /note="color: #ffff00"
     primer_bind     2330..2344
                     /label=5Gly_seq_1
                     /note="color: black; sequence: ggtggcggaggtggc"
     CDS             join(2345..2347,2348..3058)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /note="This feature has 2 segments:
                        1: 2345 .. 2347 / #05fd14 / 1a
                        2: 2348 .. 3058 / #05fd14"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(3039..3077)
                     /label=Fragment.REV2
                     /note="color: black; sequence: 
                     tggtttagtggtaacctcacttgtacagctcgtccatgc"
     promoter        complement(3303..3321)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(3339..3355)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    3363..3379
                     /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)."
                     /note="color: #31849b"
     promoter        complement(join(3387..3393,3394..3411,3412..3417))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 3387 .. 3393 / #ffffff / -10
                        2: 3394 .. 3411 / #ffffff
                        3: 3412 .. 3417 / #ffffff / -35"
     protein_bind    3432..3453
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(3741..4330)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
ORIGIN
        1 aaacaggaag gcaaatgccg aaaaaaggga ataagggcga cacggaaatg ttgaatactc
       61 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
      121 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
      181 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
      241 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
      301 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
      361 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca
      421 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
      481 agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
      541 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
      601 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgaa
      661 ttgtaatacg actcactata gggcgaatta attcgagctc ggtacccagc ttgcttgttc
      721 tttttgcaga agctcagaat aaacgctcaa ctttggcaga tcaattcccc ggtgctagat
      781 ggcggacctc agcttttctg acggcgatcc tactgtacgc acgcttttgc ggcgcgtgct
      841 ggagacggcg gattcgcgca ctccgatgag acgccggagc actcggatca atgcacaaag
      901 aagacggtct caaacgccat actctaacag gcagggtagc caaacaaaga caagtgctag
      961 gaagcaatcc catggagcta ggtcagttgg cagatcgact cgtgttcaag gtcgtgggcg
     1021 cctggaggaa cagacacccc ggactctgct aaggaatatc ttactaactg ccccagaatc
     1081 gtccactgtg atgccagacc cagtggtgaa gccagcacaa gtaccagaag tggcccggtc
     1141 ctccagacgg gagagcagtc ggggaagcct ggagctacac cttcctgaac tggagccccc
     1201 ctcaaccctg gctccgggtc taacagcccc tggcaaaagg aagcagaaac tgagattatc
     1261 agtatttcag caggaagtgg accaggggtt gcctctctcc caagagccac gtaggagtag
     1321 gagtgctgat gtttcatcct tggctagctc cttcaatctt acctttgttc tacctggtca
     1381 gccagagaca gtggagagac ctggtctggc tcgaagacgt cccatccgcc agcttgtgaa
     1441 tgctggtgct cttttgcagg atctggaaga caattcctta gcctctgctc ttccaggtga
     1501 cagtcacaga acacctgttg ctgctctgcc gatggatgta ggcttggaag acacacagcc
     1561 tttctcacag tccttggctg ccttttccct tagtgggaaa cattctctgc ccagcccttc
     1621 tcgccctggg gttgaagatg ttgaaagagt tatgggccca ccaagctcgg ggaccaggct
     1681 gcaaagccgg atgagcagat ctggccctgc tgcctctcct tcccccttcc tggagccaca
     1741 gcctccgcct gcagaaccga gagaagcagt aggatccaac gaggctgcgg agcccaagga
     1801 tcaagaagga tcttcagggt atgaggaaac ttctgctagg ccagcaagtg gagaattatc
     1861 ctccagcaca catgactcac tcccagctga acagccaccc ccgtcacctg gagttgcagt
     1921 cctatcttca gagcccctgg agtctgtgac agctaagtgc ccatccagga cccaaactgc
     1981 aggtcctagg cgccgccaag acccccacaa ggctggactg agcccctatg tgaagttctt
     2041 cagcttctgt actaagatgc ccgtggagaa gacggctctt gagatagtag agaagtgcct
     2101 agacaagtat ttccagcacc tttgcaatga cctggaggta tttgcttctc atgctggccg
     2161 taagattgtg aagccagagg acttattgct gctgatgcga cggcagggac tagtgactga
     2221 tcaagtctcg cagcatgtgc tcgtggagcg gtacctgccc ttggagtacc ggcaacaact
     2281 tatcccttgt gcattcagtg gcaattctgt cttccctgca cagccggtcg gtggcggagg
     2341 tggcgtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     2401 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     2461 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     2521 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
     2581 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     2641 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     2701 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     2761 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa
     2821 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
     2881 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     2941 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     3001 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagtg
     3061 aggttaccac taaaccagcc tcaagaacac ccgaatggag tctctaagct acataatacc
     3121 aacttacact ttacaaaatg ttgtccccca aaatgtagcc attcgtatct gctcctaata
     3181 aaaagaaagt ttcttcacat tctaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaccc
     3241 cccccccccc cccctgcagg cgcgccgccg gcgatttaaa ttaattaatc tagcttgagt
     3301 attctatagt gtcacctaaa tagcttggcg taatcatggt catagctgtt tcctgtgtga
     3361 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     3421 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     3481 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     3541 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     3601 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     3661 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     3721 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     3781 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     3841 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     3901 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     3961 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4021 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgaccttatc
     4081 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     4141 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     4201 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     4261 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     4321 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     4381 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     4441 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     4501 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     4561 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     4621 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     4681 ccagccagcc cggaagggcc gagcgcagaa gtggtcctgc aaactttatc cgcctccatc
     4741 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4801 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4861 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4921 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4981 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     5041 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     5101 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     5161 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     5221 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     5281 agcgtttctg ggtgagccct ttcaccagcg tttctgggtg agc
//
