Strain#..: ?
Box......: ?
Position.: ?
Importing the pydna package. Pydna is open source, documentated here and has a support forum as well as a publication:
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from pydna.all import *
Read the cloning vector from a local file.
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pYPKa =read("pYPKa.gb")
This vector should be a circular 3128 bp DNA molecule.
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pYPKa
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The circular seguid checksum of pYPKa should be
aV1eIrzOiCjvw01yvKkxDXHKLMk
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pYPKa.cseguid()
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Importing the restriction enzyme to be used for cloning from Biopython.
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from Bio.Restriction import AjiI
With the recognition site
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print(AjiI.site)
This enzyme should cut only once in pYPKa.
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pYPKa.cut(AjiI)
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pYPKa is digested with AjiI to form a linear DNA fragment.
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pYPKa_AjiI = pYPKa.linearize(AjiI)
The insert comes from a Genbank record. Access to Genbank is needed in order to download the template. If you execute this script, change the email address below to your own. Always tell Genbank who you are, when using the service.
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gb =Genbank("myemail@home.se")
The template is downloaded from Genbank below.
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template = gb.nucleotide("NM_001183514.1")
The template length and a representation:
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print(len(template))
print(str(template.seq))
The seguid checksum of the template should be
insertseguid
Two primers are used to amplify the insert:
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f,r =parse(""">555_RKI1_YPK_fwd
CGCGCCAAAATGGCTGCCGGTGTCCCAAA
>554_RKI1_YPK_rev
GGTCACTTTTCGGTAACTTCAACACT""", ds=False)
insert =pcr(f, r, template)
The primers anneal on the template like this.
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insert.figure()
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A recombinant plasmid is formed by ligating the insert PCR product to the linear vector.
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plasmid = (pYPKa_AjiI + insert).looped()
The plasmid sequence is rotated so that the origin is in the same position as for the cloning vector sequence.
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pYPKa_A_ScRKI1 = plasmid.synced(pYPKa)
Stamp sequence with cSEGUID checksum
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pYPKa_A_ScRKI1.stamp()
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Write sequence to a local file.
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pYPKa_A_ScRKI1.write("pYPKa_A_ScRKI1.gb")
Calculate cseguid:
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pYPKa_A_ScRKI1.cseguid()
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from pydna.all import *
pYPKa_A_ScRKI1 =read("pYPKa_A_ScRKI1.gb")
assert pYPKa_A_ScRKI1.cseguid() in pYPKa_A_ScRKI1.definition