from pydna.parsers import parse_primers
from pydna.readers import read
from pydna.design import primer_design
from pydna.amplify import pcr
from pydna.assembly import Assembly

p = { x.id: x for x in parse_primers("standard_primers.txt") }

pYPKpw = read("pYPKpw.gb")

from Bio.Restriction import EcoRV

pYPK_EcoRV = pYPKpw.linearize(EcoRV)

promoter_template   = read("pYPKa_Z_TDH3.gb")
gene_template       = read("SsXYL2.gb")
terminator_template = read("pYPKa_E_PGI.gb")

prom = pcr( p['577'], p['567'], promoter_template)

fp_tail = "tgcccactttctcactagtgacctgcagccgacAA"
rp_tail = "AAatcctgatgcgtttgtctgcacagatggCAC"

ins = primer_design(gene_template)
fp = fp_tail + ins.forward_primer
rp = rp_tail + ins.reverse_primer

print(fp.format("fasta"))
print(rp.format("fasta"))
with open("new_primers.txt", "a+") as f:
    f.write(fp.format("fasta"))
    f.write(rp.format("fasta"))

gene = pcr( fp, rp, gene_template)

term = pcr( p['568'], p['578'], terminator_template)

asm = Assembly( (pYPK_EcoRV, prom, gene, term), limit=31 )

asm

candidate = asm.assemble_circular()[0]

candidate.figure()

result = candidate.synced(pYPKpw)

product = pcr( p['577'], p['467'], result)

print(len(product))

print(len(product) - len(prom))

print(len(product) - len(gene))

product2 = pcr( p['468'], p['578'], result)

print(len(product2))

print(len(product2) - len(gene))

print(len(product2) - len(term))

result.cseguid()

result.locus = "pYPK0_tp_g_tp"
result.definition = "pYPK0_TDH3_SsXYL2_PGI"

result.stamp()

result.write("pYPK0_TDH3_SsXYL2_PGI.gb")

prom.program()

gene.program()

term.program()

import pydna
reloaded = read("pYPK0_TDH3_SsXYL2_PGI.gb")
reloaded.verify_stamp()