LOCUS       Exported                8329 bp ds-DNA     circular SYN 26-FEB-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8329)
  AUTHORS   R van Ineveld
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Feb 26, 2021 from SnapGene Viewer 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8329
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    16..279
                     /label=Tol2
     misc_feature    280..284
                     /label=Bgl2_misc_feature
                     /note="note=""Bgl2"" - 

                     280..285"
     misc_feature    286..1448
                     /label=hEF1a promoter
     misc_feature    286..304
                     /label=N-terminus_misc_feature [*]
                     /note="note=""N-terminus"" - 1698..1721"
     misc_feature    298..304
                     /label=Kozak_misc_feature
                     /note="note=""Kozak"" - 

                     1715..1721"
     primer_bind     complement(646..661)
                     /label=BP406 Turq2 van F7 into miniTol-Y-ER-C_Rv
     promoter        1254..1449
                     /label=mouse phosphoglycerate kinase 1 promoter_PGK pr...
                     /note="note=""PGK promoter"" - note=""mouse 
                     phosphoglycerate kinase 1 promoter"" - 

                     2671..3170"
     primer_bind     1357..1376
                     /label=BP308 seq vanaf EF1a
     primer_bind     complement(1431..1496)
                     /label=BP489 EF1a ipv CMV  Rv
     primer_bind     complement(1455..1520)
                     /label=BP245 prepare pLV for project T Rev
     primer_bind     1498..1528
                     /label=BP326 Primer YPet into F7 Fw
     primer_bind     1503..1571
                     /label=BP306 mVenus into F7 Fw
     primer_bind     complement(1515..1531)
                     /label=BP094 Fusion PCR PGK-Puro 02 Rev
     primer_bind     1517..1571
                     /label=BP 219 EKAREVnls to pLV Fw
     primer_bind     1517..1556
                     /label=BP 225 EKAREVnls to pLV Fw
     primer_bind     1522..1569
                     /label=BP197 Flag-mNeon into F2 Fw
     primer_bind     1533..1556
                     /label=BP 226 EKAREVnls to E4 (or EF1a) Fw
     primer_bind     complement(1542..1561)
                     /label=BP336 Seq multi-acceptor Rev
     primer_bind     1542..1561
                     /label=BP335 Seq multi-acceptor Fw
     primer_bind     1552..1571
                     /label=BP460 Scarlet-I into K19 Fw
     primer_bind     1553..1574
                     /label=BP 050 Lgr5BAC_HA1_GFP_fw
     primer_bind     1553..1572
                     /label=BP317 Make Q2 PCR2 Fw
     primer_bind     1553..1571
                     /label=BP464 Scarlet-I into K9 PCR2 Fw
     primer_bind     1553..1571
                     /label=BP 221 EKAREVnls to E4 (or EF1a) Fw
     primer_bind     1553..1571
                     /label=BP323 mVenus into F11 Fw
     primer_bind     1553..1571
                     /label=BP081 GFP in pCS2 Fw
     primer_bind     1553..1570
                     /label=BP157 mTurq2-NES into C2 Fw
     primer_bind     1553..1569
                     /label=BP193 mNeon into CFP-CX Fw
     primer_bind     1553..1569
                     /label=BP458 Scarlet-I into K4 Fw
     primer_bind     1553..1569
                     /label=BP185 mNeon into K1 Fw
     primer_bind     1553..1569
                     /label=BP195 mNeon-CAAX into K5 Fw
     primer_bind     1553..1568
                     /label=BP468 Make TV22 PCR1 Fw
                     /note="(67 bp)"
     misc_feature    1554..2264
                     /label=YPet
     primer_bind     1554..1571
                     /label=BP401 Fragm2_Fw
     primer_bind     1554..1571
                     /label=BP393 mCh-IRES to T24 Fw
     primer_bind     1554..1571
                     /label=BP201 GFP-CENP-B into K10 Fw
     primer_bind     1554..1570
                     /label=BP159 mCherry-CAAX to C8-Y+C  Fw
     primer_bind     1621..1640
                     /label=BP 010 seq GFP Rev70
     primer_bind     complement(1628..1646)
                     /label=BP 053 GFP_HA1_control_rev
     primer_bind     complement(1675..1691)
                     /label=BP 009 seq GFP Rev120
     primer_bind     complement(1681..1704)
                     /label=BP360 Make Q8c Rev
     primer_bind     2093..2112
                     /label=BP 007 seq GFP Fw540
     primer_bind     complement(2259..2278)
                     /label=BP327 Primer YPet into F7 Rev
     misc_feature    2271..2435
                     /label=WW domain
     primer_bind     complement(2389..2406)
                     /label=BP334 Seq EKAREV from WW
     primer_bind     complement(2446..2465)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2446..2465)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2448..2663
                     /label=EV linker
     misc_feature    2448..2459
                     /label=Feature 23
     primer_bind     complement(2458..2483)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2458..2483)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2460..2471
                     /label=Feature 24
     misc_feature    2472..2477
                     /label=Feature 25
     misc_feature    2478..2489
                     /label=Feature 43
     primer_bind     complement(2488..2507)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2488..2507)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2490..2501
                     /label=Feature 32
     primer_bind     complement(2500..2525)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2500..2525)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2502..2519
                     /label=Feature 46
     primer_bind     complement(2518..2537)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2518..2537)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2520..2531
                     /label=Feature 33
     primer_bind     complement(2530..2549)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2530..2549)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2532..2543
                     /label=Feature 34
     primer_bind     complement(2542..2561)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2542..2561)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2544..2555
                     /label=Feature 35
     primer_bind     complement(2554..2579)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2554..2579)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2556..2567
                     /label=Feature 36
     misc_feature    2568..2585
                     /label=Feature 44
     primer_bind     complement(2584..2603)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2584..2603)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2586..2597
                     /label=Feature 37
     primer_bind     complement(2596..2621)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2596..2621)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2598..2615
                     /label=Feature 47
     primer_bind     complement(2614..2633)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2614..2633)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2616..2627
                     /label=Feature 38
     misc_feature    2628..2639
                     /label=Feature 39
     misc_feature    2640..2651
                     /label=Feature 40
     primer_bind     2650..2699
                     /label=BP374 Mut F7 S413P+G414S Fw?????????
     primer_bind     2650..2699
                     /label=BP372 Mut F7 G414S Fw?????????
     primer_bind     complement(2650..2699)
                     /label=BP375 Mut F7 S413P+G414S Rev????????
     primer_bind     complement(2650..2699)
                     /label=BP373 Mut F7 G414S Rev????????
     misc_feature    2652..2663
                     /label=Feature 41
     primer_bind     complement(2668..2729)
                     /label=BP414 Mut F7 D423K, R419S Rv
     primer_bind     2668..2729
                     /label=BP413 Mut F7 D423K,R419S Fw
     primer_bind     complement(2671..2710)
                     /label=BP392 Mut F7-PW TA Rv
     primer_bind     2671..2710
                     /label=BP391 Mut F7-PW TA Fw
     primer_bind     complement(2671..2702)
                     /label=BP357 F7-P-mimetic E Rev
     primer_bind     complement(2671..2702)
                     /label=BP355 F7-P-mimetic D Rev
     primer_bind     complement(2673..2729)
                     /label=BP377 Mut F7 V422T Rv
     primer_bind     2673..2729
                     /label=BP376 Mut F7 V422T Fw
     misc_feature    2676..2729
                     /label=ERK Substrate
     primer_bind     complement(2676..2729)
                     /label=BP412 Mut F7 D423K Rv
     primer_bind     2676..2729
                     /label=BP411 Mut F7 D423K Fw
     primer_bind     complement(2677..2728)
                     /label=BP408 Mut F7 PW-to-W Rv
     primer_bind     2677..2728
                     /label=BP407 Mut F7 PW-to-W Fw
     primer_bind     complement(2679..2733)
                     /label=BP386 Mut F7 KK424,426PE Rv
     primer_bind     2679..2733
                     /label=BP385 Mut F7 KK424,426PE Fw
     primer_bind     complement(2679..2733)
                     /label=BP384 Mut F7 K426E Rv
     primer_bind     2679..2733
                     /label=BP383 Mut F7 K426E Fw
     primer_bind     complement(2679..2733)
                     /label=BP382 Mut F7 K426W Rv
     primer_bind     2679..2733
                     /label=BP381 Mut F7 K426W Fw
     primer_bind     complement(2679..2733)
                     /label=BP380 Mut F7 KK424,426PW Rv
     primer_bind     2679..2733
                     /label=BP379 Mut F7 KK424,426PW Fw
     primer_bind     complement(2679..2733)
                     /label=BP371 Mut F7 KK424,426PP Rv
     primer_bind     2679..2733
                     /label=BP370 Mut F7 KK424,426PP Fw
     primer_bind     complement(2679..2728)
                     /label=BP367 Mut F7 K424P Rev??????
     primer_bind     2679..2728
                     /label=BP366 Mut F7 K424P Fw?????? ???????????????
     primer_bind     complement(2682..2733)
                     /label=BP369 Mut F7 K426P Rev????????
     primer_bind     2682..2733
                     /label=BP368 Mut F7 K426P Fw?????????
     primer_bind     complement(2685..2733)
                     /label=BP410 Mut F7 PW-to-P Rv
     primer_bind     2685..2733
                     /label=BP409 Mut F7 PW-to-P Fw
     primer_bind     2689..2738
                     /label=BP435 Mut F7 Dock F429D Fw
     misc_feature    2691..2693
                     /label=fosf.site
     primer_bind     2694..2744
                     /label=BP415 Mut F7 No Dock Fw
     CDS             2703..2705
                     /label=P
     CDS             2709..2711
                     /label=W
     primer_bind     2733..2752
                     /label=BP332 Primer Turq2 into F20 Fw
     primer_bind     2733..2752
                     /label=BP364 Make F22 Fw
     primer_bind     2738..2762
                     /label=BP421 Turq2 to YERC Frgm2 Fw
     CDS             2739..3449
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)Mutation K206A = monomeric to sticky
                     Mutation V224L = enhances dimerization sysynergistically 
                     with S208F of YPet (Vinkenborg. J.L. et al., 2007)
                     "
                     /label=Turquoise2 FRET-optimized
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFLTAAGITLGMDEL"
     primer_bind     2769..2793
                     /label=BP405 Turq2 van F7 into miniTol-Y-ER-C_Fw
     variation       2934..2936
                     /label=T65S , mTurq
     variation       2937..2939
                     /label=Y66W
     variation       2955..2957
                     /label=S72A
     variation       3126..3128
                     /label=G129D
     misc_feature    3177..3179
                     /label=I146F , mTurq2
     variation       3198..3200
                     /label=M153T
     variation       3228..3230
                     /label=V163A
     primer_bind     complement(3255..3272)
                     /label=BP363 Seq Rev vanaf mutated Turq
     primer_bind     3255..3272
                     /label=BP345 Seq Fw vanaf mutated Turq
     variation       3264..3266
                     /label=S175G
     misc_feature    3357..3359
                     /label=K206A monomeric->sticky
     variation       3411..3413
                     /label=V224L enhances dimerization
     primer_bind     complement(3432..3450)
                     /label=BP422 Turq2 to YERC Frgm2 Rev
     misc_feature    3450..3454
                     /label=eCFP
     CDS             3456..3476
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3467..3499)
                     /label=BP 220 EKAREVnls to pLV Rev
     primer_bind     complement(3467..3489)
                     /label=BP 222 EKAREVnls to E4 (or EF1a) Rev
     primer_bind     complement(3507..3522)
                     /label=BP454 H2B-Neon to F80 Rv
     primer_bind     3532..3558
                     /label=BP295 make S1, polyA into E4, Fw
     promoter        complement(3532..3549)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3543..3574
                     /label=BP499 polyA into TV22 Fw
     polyA_signal    3554..3688
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3570..3596)
                     /label=BP456 Insulator into T30 Fw
     primer_bind     complement(3570..3596)
                     /label=BP466 pUNKI-insulator into K4 Fw
     primer_bind     3631..3654
                     /label=BP190 Seq Rev into TagGFP of Casper3
     primer_bind     complement(3728..3753)
                     /label=BP 043 PGK-Puro-pA Rv
     primer_bind     complement(3728..3753)
                     /label=BP077 PGK-Puro-pA Rv
     primer_bind     complement(3728..3753)
                     /label=BP500 polyA into TV22 Rv
     primer_bind     complement(3729..3754)
                     /label=BP475 Make TV26 PCR2 Rev
     primer_bind     complement(3730..3771)
                     /label=BP296 make S1, polyA into E4, Rev
     primer_bind     3730..3749
                     /label=BP067 Seq primer polyA Fw
     primer_bind     complement(3730..3749)
                     /label=BP066 Seq primer polyA Rev
     primer_bind     complement(3733..3762)
                     /label=bp341 TagRFP into Q3 Rev
     primer_bind     complement(3738..3759)
                     /label=BP439 Make F7-NES Rev
     primer_bind     complement(3743..3771)
                     /label=BP333 Primer Turq2 into F20 Rev
     primer_bind     3746..3778
                     /label=BP 042 PGK-Puro-pA Fw
     primer_bind     3746..3765
                     /label=BP462 Scarlet-I into K9 PCR1 Fw
     primer_bind     3756..3772
                     /label=BP447 2xPCR DeathStar_2 Rv
     misc_feature    3757..3771
                     /label=Infusion homology arm
     primer_bind     3763..3778
                     /label=BP169 PGK:Puro w/o polyAis Fw
     primer_bind     3763..3778
                     /label=BP167 PGK:Puro w/o polyAis Fw
     primer_bind     3765..3780
                     /label=BP229 PGK-PuroDTK for T2 Fw
     primer_bind     3765..3780
                     /label=BP249 PGK-PruDTK for T2 Fw
     promoter        3766..4265
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3805..3821)
                     /label=BP406 Turq2 van F7 into miniTol-Y-ER-C_Rv
     primer_bind     3918..3936
                     /label=BP119 PGK Seq Fw170
     primer_bind     complement(4167..4177)
                     /label=BP435 Mut F7 Dock F429D Fw
     primer_bind     complement(4198..4217)
                     /label=BP120 PGK Seq Rev 430 (2)
     primer_bind     complement(4198..4217)
                     /label=BP120 PGK Seq Rev 430
     primer_bind     complement(4266..4285)
                     /label=BP484 CreER into TV26-EcoRI Rv
     primer_bind     complement(4266..4285)
                     /label=BP482 CreER into TV26 Rv
     promoter        4277..4324
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     primer_bind     complement(4307..4334)
                     /label=BP240 prepare pLV for project T Rev
     primer_bind     complement(4307..4334)
                     /label=BP242 prepare pLV for project T Rev
     primer_bind     4337..4384
                     /label=BP040 Puro Fw
     primer_bind     4343..4362
                     /label=BP326 Primer YPet into F7 Fw
     primer_bind     complement(4349..4418)
                     /label=BP094 Fusion PCR PGK-Puro 02 Rev
     primer_bind     4367..4392
                     /label=BP095 Fusion PCR PGK-Puro 03 Fw
     CDS             4373..4972
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4508..4527)
                     /label=BP123 Puro Seq Rev140
     primer_bind     complement(4772..4789)
                     /label=BP124 Puro Seq Rev410
     primer_bind     4772..4789
                     /label=BP121 Puro Seq Fw400
     primer_bind     4869..4888
                     /label=BP122 Puro Seq Fw510
     primer_bind     complement(4956..4987)
                     /label=BP098 Fusion PCR PGK-Puro 06 Rev
     primer_bind     complement(4956..4987)
                     /label=BP041 Puro Rv
     primer_bind     complement(4956..4973)
                     /label=BP234 PGK-Puro into T4 Rev
     primer_bind     complement(4956..4972)
                     /label=BP096 Fusion PCR PGK-Puro 04 Rev
     primer_bind     complement(4956..4972)
                     /label=BP170 PGK:Puro w/o polyAis Rev
     primer_bind     complement(4956..4972)
                     /label=BP168 PGK:Puro w/o polyAis Rev
     primer_bind     complement(4956..4972)
                     /label=BP471 Make TV22 PCR2 Rv
     primer_bind     4979..5005
                     /label=BP295 make S1, polyA into E4, Fw
     promoter        complement(4979..4996)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     4990..5021
                     /label=BP499 polyA into TV22 Fw
     polyA_signal    5001..5135
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5017..5043)
                     /label=BP456 Insulator into T30 Fw
     primer_bind     complement(5017..5043)
                     /label=BP466 pUNKI-insulator into K4 Fw
     primer_bind     5078..5101
                     /label=BP190 Seq Rev into TagGFP of Casper3
     primer_bind     5138..5169
                     /label=BP474 Make TV26 PCR2 Fw
     primer_bind     5139..5169
                     /label=BP470 Make TV22 PCR2 Fw
     primer_bind     complement(5175..5206)
                     /label=BP077 PGK-Puro-pA Rv
     primer_bind     complement(5175..5200)
                     /label=BP 043 PGK-Puro-pA Rv
     primer_bind     complement(5175..5200)
                     /label=BP500 polyA into TV22 Rv
     primer_bind     complement(5176..5200)
                     /label=BP475 Make TV26 PCR2 Rev
     primer_bind     complement(5177..5200)
                     /label=BP296 make S1, polyA into E4, Rev
     primer_bind     5177..5196
                     /label=BP067 Seq primer polyA Fw
     primer_bind     complement(5177..5196)
                     /label=BP066 Seq primer polyA Rev
     primer_bind     complement(5180..5200)
                     /label=bp341 TagRFP into Q3 Rev
     primer_bind     complement(5185..5200)
                     /label=BP439 Make F7-NES Rev
     misc_feature    5207..5418
                     /label=Tol2
     primer_bind     5428..5445
                     /label=BP449 adapt miniTol for INS Rv
     promoter        complement(5454..5472)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5490..5506)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5514..5530
                     /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(5538..5568)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5565..5584)
                     /label=BP258 Seq pr in miniTol vector (T2) Rev
     protein_bind    5583..5604
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5892..6480)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6651..7511)
                     /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     complement(6666..6685)
                     /label=BP 048 Amp_HA2_control_fw
     promoter        complement(7512..7616)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7694..8149)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     8290..8306
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        join(8313..8329,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     join(8325..8329,1..15)
                     /label=BP213 Mash3x into E7-gcamp Fw
ORIGIN
        1 gggcgaattg ggcccagagg tgtaaagtac ttgagtaatt ttacttgatt actgtactta
       61 agtattattt ttggggattt ttactttact tgagtacaat taaaaatcaa tacttttact
      121 tttacttaat tacatttttt tagaaaaaaa agtacttttt actccttaca attttattta
      181 cagtcaaaaa gtacttattt tttggagatc acttcattct attttccctt gctattacca
      241 aaccaattga attgcgctga tgcccagttt aatttaaata gatctggctc cggtgcccgt
      301 cagtgggcag agcgcacatc gcccacagtc cccgagaagt tggggggagg ggtcggcaat
      361 tgaaccggtg cctagagaag gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg
      421 ctccgccttt ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac
      481 gttctttttc gcaacgggtt tgccgccaga acacaggtaa gtgccgtgtg tggttcccgc
      541 gggcctggcc tctttacggg ttatggccct tgcgtgcctt gaattacttc cacctggctc
      601 cagtacgtga ttcttgatcc cgagctggag ccaggggcgg gccttgcgct ttaggagccc
      661 cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg gggccgccgc gtgcgaatct
      721 ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc tctagccatt taaaattttt
      781 gatgacctgc tgcgacgctt tttttctggc aagatagtct tgtaaatgcg ggccaggatc
      841 tgcacactgg tatttcggtt tttggggccg cgggcggcga cggggcccgt gcgtcccagc
      901 gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc gagaatcgga cgggggtagt
      961 ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc gccgtgtatc gccccgccct
     1021 gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc ggaaagatgg ccgcttcccg
     1081 gccctgctcc agggggctca aaatggagga cgcggcgctc gggagagcgg gcgggtgagt
     1141 cacccacaca aaggaaaggg gcctttccgt cctcagccgt cgcttcatgt gactccacgg
     1201 agtaccgggc gccgtccagg cacctcgatt agttctggag cttttggagt acgtcgtctt
     1261 taggttgggg ggaggggttt tatgcgatgg agtttcccca cactgagtgg gtggagactg
     1321 aagttaggcc agcttggcac ttgatgtaat tctccttgga atttgccctt tttgagtttg
     1381 gatcttggtt cattctcaag cctcagacag tggttcaaag tttttttctt ccatttcagg
     1441 tgtcgtgagg atcccccggg ctgcaggaat tcggcgcgcc acgcgtaccg atagcttcga
     1501 attctgcagt cgacgatatc actagtctcg agcattttgg caaagaattc ggcatggtga
     1561 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     1621 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
     1681 tgaccctgaa gcttctatgc accaccggca agctgcccgt gccctggccc accctcgtga
     1741 ccaccctggg ctacggcctg cagtgcttcg cccgctaccc cgaccacatg aagcagcacg
     1801 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     1861 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     1921 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     1981 agtacaacta caacagccac aacgtctata tcaccgccga caagcagaag aacggcatca
     2041 aggccaactt caagatccgc cacaacatcg aggacggcgg cgtgcagctc gccgaccact
     2101 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     2161 gctaccagtc cgccctgttc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     2221 agttcctgac cgccgccggg atcactgagg gcatgaacga gctgtacctc gagatggcgg
     2281 acgaggagaa gctgccgccc ggctgggaga agcgcatgag ccgcagctca ggccgagtgt
     2341 actacttcaa ccacatcact aacgccagcc agtgggagcg gcccagcggc aacagcagca
     2401 gtggtggcaa aaacgggcag ggggagcctg ccaggggtac cagtgctggt ggtagtgctg
     2461 gtggtagtgc tggtggttcc ggcagtgctg gtggtagtgc tggtggtagt accagtgctg
     2521 gtggtagtgc tggtggtagt gctggtggta gtgctggtgg tagtgctggt ggttccggca
     2581 gtgctggtgg tagtgctggt ggtagtacca gtgctggtgg tagtgctggt ggtagtgctg
     2641 gtggtagtgc tggtggtagt gctggtggtt ccggaccaga tgtccctaga actccagtgg
     2701 atccagcatg gctgtcattc caatttccgg gcggccgcat ggtgagtaaa ggggaagaac
     2761 tgtttactgg cgtggtgcca attctggtcg aactggatgg agatgtaaat gggcataaat
     2821 ttagtgtgtc tggagaaggg gaaggagacg ctacatatgg gaaactgaca ctgaaattta
     2881 tttgtactac tggaaaactg cctgtgcctt ggcctacact cgtgacaacc ctgtcttggg
     2941 gagtgcagtg ttttgcccgg tatcctgatc atatgaaaca gcatgatttt ttcaaatctg
     3001 ccatgcctga gggatatgtc caggaacgga caattttctt taaagatgat gggaattata
     3061 aaacacgggc cgaagtgaaa tttgaagggg atacactggt gaatcggatt gaactgaaag
     3121 gaattgattt taaagaagat gggaatattc tgggccataa actggaatat aattatttca
     3181 gcgataatgt ctacattaca gccgataaac agaagaatgg gattaaagcc aattttaaaa
     3241 ttcggcataa tattgaagat ggaggggtgc agctcgccga tcattatcag cagaatacac
     3301 ctattgggga tgggcctgtg ctgctgcctg ataatcatta tctgagcaca cagtctgccc
     3361 tgagcaagga ccctaatgaa aagcgggacc atatggtcct gctggaattt ctgactgccg
     3421 ccggcattac actcgggatg gatgaactgt ctagacctaa gaagaagcgt aaggtgtaag
     3481 tcgacggtcg agtcgagacg ctagccaccg gtcgttaaca ttaagctaga actatagtga
     3541 gtcgtattac gtagatccag acatgataag atacattgat gagtttggac aaaccacaac
     3601 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     3661 aaccattata agctgcaata aacaagttaa caacgccggc catcgatatg tttaaactca
     3721 ggttcagggg gaggtgtggg aggtttttta attcgcggcc gcaccgggta ggggaggcgc
     3781 ttttcccaag gcagtctgga gcatgcgctt tagcagcccc gctgggcact tggcgctaca
     3841 caagtggcct ctggcctcgc acacattcca catccaccgg taggcgccaa ccggctccgt
     3901 tctttggtgg ccccttcgcg ccaccttcta ctcctcccct agtcaggaag ttcccccccg
     3961 ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc actagtctcg
     4021 tgcagatgga cagcaccgct gagcaatgga agcgggtagg cctttggggc agcggccaat
     4081 agcagctttg ctccttcgct ttctgggctc agaggctggg aaggggtggg tccgggggcg
     4141 ggctcagggg cgggctcagg ggcggggcgg gcgcccgaag gtcctccgga ggcccggcat
     4201 tctgcacgct tcaaaagcgc acgtctgccg cgctgttctc ctcttcctca tctccgggcc
     4261 tttcgacctg cagcctgttg acaattaatc atcggcatag tatatcggca tagtataata
     4321 cgacaaggtg aggaactaaa ccgtcgacga tttcactagt ctcgaagcca ccatgaccga
     4381 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     4441 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     4501 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     4561 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     4621 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     4681 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     4741 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     4801 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
     4861 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     4921 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gactagaact
     4981 atagtgagtc gtattacgta gatccagaca tgataagata cattgatgag tttggacaaa
     5041 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     5101 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     5161 tgtttcaggt tcagggggag gtgtgggagg ttttttaatt aagcttaaac aagaatctct
     5221 agttttcttt cttgctttta cttttacttc cttaatactc aagtacaatt ttaatggagt
     5281 acttttttac ttttactcaa gtaagattct agccagatac ttttactttt aattgagtaa
     5341 aattttccct aagtacttgt actttcactt gagtaaaatt tttgagtact ttttacacct
     5401 ctgctcgacc atatgggaga gctcccaacg cgttggatgc atagcttgag tattctatag
     5461 tgtcacctaa atagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     5521 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     5581 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     5641 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     5701 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     5761 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     5821 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     5881 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5941 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     6001 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     6061 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     6121 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     6181 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     6241 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     6301 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     6361 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     6421 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     6481 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     6541 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     6601 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     6661 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     6721 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     6781 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     6841 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     6901 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     6961 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     7021 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     7081 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     7141 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     7201 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     7261 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     7321 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     7381 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     7441 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     7501 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     7561 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     7621 tttccccgaa aagtgccacc tgatgcggtg tgaaataccg cacagatgcg taaggagaaa
     7681 ataccgcatc aggaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt
     7741 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa
     7801 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag
     7861 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt
     7921 gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac
     7981 cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag
     8041 gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg
     8101 cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc cattcgccat
     8161 tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc
     8221 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     8281 cacgacgttg taaaacgacg gccagtgaat tgtaatacga ctcactata
//
