Composite

Part:BBa_K277135

Designed by: James DiCarlo   Group: iGEM09_Johns_Hopkins-BAG   (2009-10-21)


3L.3_23.A2


3L.3_23.A2 was designed as a piece of synthetic chromosome 3 with the goal of minimizing and stabilizing that chromosome and to that end has had any tRNAs, introns,repeat regions, and transposons that were present in the wildtype chromosome removed. In addition a very few gene sequences were slightly recoded to add or remove restriction enzyme recognition sites to facilitate assembly; most gene sequences were slightly recoded to introduce unique primers for diagnostic PCR amplification, and some gene sequences were slightly recoded to address the distribution of stop codon usage. 3L.3_23.A2 is comprised of and must be constructed from the parts 3L.3_23.A2.01, 3L.3_23.A2.02, 3L.3_23.A2.03, 3L.3_23.A2.04, 3L.3_23.A2.05, 3L.3_23.A2.06, 3L.3_23.A2.07, 3L.3_23.A2.08, 3L.3_23.A2.09, 3L.3_23.A2.10, 3L.3_23.A2.11, 3L.3_23.A2.12, 3L.3_23.A2.13, and 3L.3_23.A2.14.

This part wholly contains the following features (positions offset from first base of sequence):

kind and name offset notes

reverse_primer YCL057W_tagr1v1 (6353..6380)

loxP_site loxPsym_YCL057W (8175..8208)

forward_primer YCL059C_tagf1v1 (3862..3889)

forward_primer YCL057W_tagf1v1 (6158..6185)

reverse_primer YCL055W_tagr1v1 (9956..9983)

gene YCL056C (8224..8658) Protein of unknown function%3B green fluorescent protein (GFP)-fusion protein localizes to the cytoplasm in a punctate pattern

loxP_site loxPsym_YCL055W (10239..10272)

gene YCL059C (3694..4644) Essential nucleolar protein required for the synthesis of 18S rRNA and for the assembly of 40S ribosomal subunit

mutation_affecting_coding_sequence YCL057W_re_remove_AlwNI (6474..6485) removal of AlwNI

mutation_affecting_coding_sequence YCL061C_re_remove_AlwNI (1548..1559) removal of AlwNI

reverse_primer YCL061C_tagr3v1 (3219..3246)

gene YCL058W-A (4849..5190) Protein of unknown function%3B identified by homology to Ashbya gossypii

reverse_primer YCL061C_tagr2v1 (2808..2835)

reverse_primer YCL061C_tagr1v1 (2601..2628)

reverse_primer YCL057W_tagr2v1 (7577..7604)

forward_primer YCL061C_tagf3v1 (2991..3018)

loxP_site loxPsym_YCL061C (44..77)

gene YCL055W (9228..10235) Transcription factor required for induction of KAR3 and CIK1 during mating%2C also required during meiosis%3B exists in two forms%2C a slower-migrating form more abundant during vegetative growth and a faster-migrating form induced by pheromone

forward_primer YCL057W_tagf2v1 (7181..7208)

gene YCL057W (6033..8171) Zinc metalloendopeptidase%2C found in the cytoplasm and intermembrane space of mitochondria%3B with Cym1p%2C involved in degradation of mitochondrial proteins and of presequence peptides cleaved from imported proteins

gene YCL061C (81..3371) S-phase checkpoint protein found at replication forks%2C required for DNA replication%3B also required for Rad53p activation during DNA replication stress%2C where it forms a replication-pausing complex with Tof1p and is phosphorylated by Mec1p%3B protein involved in replication checkpoint

forward_primer YCL061C_tagf1v1 (2208..2235)

mutation_affecting_coding_sequence YCL058C_stop_recode_overlap_from_YCL058W-A (5190..5190) Overlaps stop codon change in YCL058W-A

gene YCL057C-A (5297..5590) Hypothetical protein%2C has similarity to proteins in S. pombe%2C C. elegans%2C D. melanogaster.

forward_primer YCL055W_tagf1v1 (9500..9527)

reverse_primer YCL059C_tagr1v1 (4312..4336)

gene YCL058C (4788..5246) Protein of unknown function%2C required for survival upon exposure to K1 killer toxin%3B involved in ion homeostasis

forward_primer YCL061C_tagf2v1 (2337..2364)

Sequence (corresponding to coordinates 8871..19174 in synthetic chromosome yeast_chr3_3_23)

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 3146
    Illegal XbaI site found at 1426
    Illegal XbaI site found at 4476
    Illegal XbaI site found at 4503
    Illegal XbaI site found at 9454
    Illegal SpeI site found at 376
    Illegal SpeI site found at 2712
    Illegal SpeI site found at 6193
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 3146
    Illegal NheI site found at 1950
    Illegal NheI site found at 2226
    Illegal NheI site found at 4372
    Illegal NheI site found at 6436
    Illegal SpeI site found at 376
    Illegal SpeI site found at 2712
    Illegal SpeI site found at 6193
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 3146
    Illegal BglII site found at 6662
    Illegal BamHI site found at 8107
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 3146
    Illegal XbaI site found at 1426
    Illegal XbaI site found at 4476
    Illegal XbaI site found at 4503
    Illegal XbaI site found at 9454
    Illegal SpeI site found at 376
    Illegal SpeI site found at 2712
    Illegal SpeI site found at 6193
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 3146
    Illegal XbaI site found at 1426
    Illegal XbaI site found at 4476
    Illegal XbaI site found at 4503
    Illegal XbaI site found at 9454
    Illegal SpeI site found at 376
    Illegal SpeI site found at 2712
    Illegal SpeI site found at 6193
    Illegal AgeI site found at 3256
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 2922
    Illegal SapI site found at 3099


[edit]
Categories
Parameters
n/a 3L.3_23.A2