pYPK3

pYPK3 is a version of pYPK1 that has a CEN6/ARS yeast ori and a HphMX4 marker instead of the bleomycin marker.



In [1]:

    
from pydna.all import *
pYPK1 =read("pYPK1.gb")
gb =Genbank("bjornjobb@gmail.com")



In [2]:

    
githubuser="BjornFJohansson"
gistid = "c5424b7ebbf553c52053"
x=download_text('https://gist.githubusercontent.com/{user}/{gistid}/raw'.format(user=githubuser, gistid=gistid))
pAG32 =read(x)



In [3]:

    
pAG32









    Out[3]:





Dseqrecord(o4160)

pAG32 sequence



In [4]:

    
p150,p149,p325,p324,p678,p666 = parse_primers('''
>150_MX4fwd (25-mer) Primer in the Ashbya gossypi TEF promoter in forward direction. contaminated?
AAAATCTTGCTAGGATACAGTTCTC

>149_MX4rev (25-mer) Primer in the Ashbya gossypi TEF terminator in the reverse direction.
ACAAATGACAAGTTCTTGAAAACAA

>325_Hygfwd (27-mer)
GATGTAGGAGGGCGTGGATATGTCCTG

>324_Hygrev (27-mer)
TGCATCATCGAAATTGCCGTCAACCAA

>678_pYPK0_hygfwd: (77-mer)
ctcacgttaagggattttggtcatgagCACATACGATTTAGGTGACACTATAGAAC

>666_pYPK0_hygrev (70-mer)
catctttgacagcttatcatcgataagctCGACTCACTATAGGGAGACC''')



In [5]:

    
hygmarker1 =pcr( p678, p324, pAG32 )



In [6]:

    
hygmarker2 =pcr( p325, p666, pAG32 )



In [7]:

    
from Bio.Restriction import PvuII



In [8]:

    
vect = pYPK1.linearize(PvuII)



In [9]:

    
a=Assembly([hygmarker1, hygmarker2, vect], limit =251)
a









    Out[9]:





Assembly
fragments..: 1371bp 1262bp 4640bp
limit(bp)..: 251
G.nodes....: 6
algorithm..: common_sub_strings



In [10]:

    
candidate = a.assemble_circular()[0]



In [11]:

    
candidate.figure()









    Out[11]:





 -|1371bp_PCR_prod|741
|                  \/
|                  /\
|                  741|1262bp_PCR_prod|251
|                                      \/
|                                      /\
|                                      251|pYPK1_lin_rc|373
|                                                       \/
|                                                       /\
|                                                       373-
|                                                          |
 ----------------------------------------------------------



In [12]:

    
pYPK3=candidate.synced(pYPK1)



In [13]:

    
pYPK3.stamp()









    Out[13]:





cSEGUID_HGdQuJehFgtyDtEoXeOty-J5sg8



In [14]:

    
pYPK3.locus = "pYPK3"



In [15]:

    
pYPK3.write("pYPK3.gb")









    




pYPK3.gb

DOWNLOAD



In [16]:

    
from pydna.all import *
reloaded=read("pYPK3.gb")
assert reloaded.cseguid() in reloaded.definition