如何对构建基因克隆的测序结果批量进行序列比对并输出结果

   做过克隆构建的人都知道，有时候我们需要对测序结果进行分析，查看测序结果是否与我们想要构建的序列是否一致，即使是构建同一条序列的克隆，有时候根据实验实际情况，我们需要测序的单克隆不止两三个，有可能是五六个，也有可能是几十上百个，面对大量的测序结果，如果我们进行一一翻译比对，那么无疑花费我们大量时间，耗费不少精力，下面展示如何通过代码进行批量的处理并输出结果。

一、读取测序数据并进行相关设置

import os
path = "E:\gene_counts"
path1 = "E:\gene_counts"
head = "CCATGG"  #输入起始翻译位置
tail = "TIEWA"         #输入目的序列尾部氨基酸
klyk = "MVQKSLLFSLLASTALGALTKRYSFPLPESQGSETFSEPYEVAAGETFDGGMKTYGRGVECTGQVEGGEDDTVFIVQEGGTLKNAIIGTDQIEGVYCLGSCTIENVWWEKVCEDALSLKEGDGIYTISGGGAQGAEDKVIQHNTGGEVIIDGFEVYDFGKLYRSCGTCGDIQRKVSVSNVVAVSGSQLVGINENFGDTATIDSSVCATDVNDICATYNGTDGDEEPEEVSTGPSDYCIYTEPIAECA"
#输入想要比对的目的序列

二、将合格测序数据整理打包

  正向（5‘端到3‘端测序）测序数据处理：

for a in os.walk(path):
  a = a[2]
​
f1 = open(path1 + "\测序结果生成.txt", "w")
for b in a:
  if b[-8:] == "[T7].seq":
    f = open(path + "/" + b, "r")
    for c in f.readlines():
      d = c
      if  head in d:
        e = d.index(head)
        m = d[e + 2:]
        f1.write(">" + b[21:-10] + "\n" + m + "\n")
        f.close()
​
f1.write(">" + "\n")
f1.close()
​

  反向（3‘端到5‘端测序）测序数据处理：

​
import os
for a in os.walk(path):
  a = a[2]
​
f1 = open(path1 + "/测序结果生成.txt", "w")
for b in a:
  if b[-12:] == "[T7-TER].seq":
    f = open(path + "/" + b, "r")
    for c in f.readlines():
      d = c
      if  head in d:
        e = d.index(head)
        m = d[0:e + 4]
        f1.write(">" + b[21:-14] + "\n" + m + "\n")
        f.close()
​
f1.write(">" + "\n")
f1.close()

正向测序结果和反向测序数据拼接：

import os
for a in os.walk(path):
  a = a[2]
​
​
zheng = {}
for b in a:
  if b[-8:] == "[T7].seq":
    f = open(path + "/" + b, "r")
    for c in f.readlines():
      d = c
      if  head in d:
        e = d.index(head)
        m = d[e + 2:]
        zheng.update({">" + b[21:-10]:m})
        f.close()
​
fan = {}
for b in a:
  if b[-12:] == "[T7-TER].seq":
    f = open(path + "/" + b, "r")
    for c in f.readlines():
      d = c
      if  head in d:
        e = d.index(head)
        m = d[0:e + 4]
        mw = ""
        for n in m:
          if n == "A":
            n = "T"
            mw += n
          elif n == "T":
            n = "A"
            mw += n
          elif n == "G":
            n = "C"
            mw += n
          elif n == "C":
            n = "G"
            mw += n
        fan.update({">" + b[21:-14]:mw[::-1]})
        f.close()
​
f1 = open(path1 + "/测序结果生成.txt", "w")
for key in zheng:
  if key in fan:
    kk = zheng[key][-200:-190]
    if kk in fan[key]:
      mm = fan[key][fan[key].index(kk):]
      yy = zheng[key][0:-200]
      nn = yy + mm
      f1.write(key + "\n" + nn + "\n")
​
f1.write(">" + "\n")
f1.close()
​

三、将翻译的数据进行清洗打包

#在线网站进行一键翻译https://www.novopro.cn/tools/translate.html
f = open(path1 + "/测序结果生成.txt", "r")
b = ""
diction1 = []
diction2 = []
i = 0
for a in f.readlines():
  i += 1
  if i == 1:
    name = a[5:-1]
  else:
    if a[0] != ">":
      b += a.replace("\n", "")
    if a[0] == ">":
      if tail in b:
        diction1.append((name,b))
        diction2.append((name,b[0:len(ClyC) + 9]))
        print((name,b[0:(len(ClyC) + 9)]))
        b = ""
        name = a[5:-1]
​
f.close()
​

四、寻找并输出突变位点（不突变输出None）

for x, y in diction2:
  if tail in y:
    if len(y[0:(y.index(tail) + len(tail))]) == len(ClyC):
      print(y)
      if y[0:(y.index(tail) + len(tail))] == ClyC:
        print(x, "None")
      else:
        for b in range(len(ClyC)):
          if ClyC[b] != y[b]:
            print(x, ":", ClyC[b] + str(b + 1) + y[b])
    elif len(y[0:(y.index(tail) + len(tail))]) > len(ClyC):
      print(y + "\n" + x , "：插入突变")
    elif len(y[0:(y.index(tail) + len(tail))]) < len(ClyC):
      print(y + "\n" + x , "：缺失突变")
​

五、输出结果展示

MVQKSLLFSLLASTALGALTKRYSFPLPESQGSETFSEPYEVAAGETFDGGMKTYGRGVECTGQVEGGEDDTVFIVQEGGTLKNAIIGTDQIEGVYCLGSCTIENVWWEKVCEDALSLKEGDGIYTISGGGAQGAEDKVIQHNTGGEVIIDGFEVYDFGKLYRSCGTCGDIQRKVSVSNVVAVSGSQLVGINENFGDTATIDSSVCATDVNDICATYNGTDGDEEPEEVSTGPSDYCIYTEPIAECA
3-3 : P81K
3-3 : S170I
MVQKSLLFSLLASTALGALTKRYSFPLPESQGSETFSEPYEVAAGETFDGGMKTYGRGVECTGQVEGGEDDTVFIVQEGGTLKNAIIGTDQIEGVYCLGSCTIENVWWEKVCEDALSLKEGDGIYTISGGGAQGAEDKVIQHNTGGEVIIDGFEVYDFGKLYRSCGTCGDIQRKVSVSNVVAVSGSQLVGINENFGDTATIDSSVCATDVNDICATYNGTDGDEEPEEVSTGPSDYCIYTEPIAECA
7-3 : Y209K
MVQKSLLFSLLASTALGALTKRYSFPLPESQGSETFSEPYEVAAGETFDGGMKTYGRGVECTGQVEGGEDDTVFIVQEGGTLKNAIIGTDQIEGVYCLGSCTIENVWWEKVCEDALSLKEGDGIYTISGGGAQGAEDKVIQHNTGGEVIIDGFEVYDFGKLYRSCGTCGDIQRKVSVSNVVAVSGSQLVGINENFGDTATIDSSVCATDVNDICATYNGTDGDEEPEEVSTGPSDYCIYTEPIAECA
9-3 : W19C
9-3 : W177L
9-3 : T251K
MVQKSLLFSLLASTALGALTKRYSFPLPESQGSETFSEPYEVAAGETFDGGMKTYGRGVECTGQVEGGEDDTVFIVQEGGTLKNAIIGTDQIEGVYCLGSCTIENVWWEKVCEDALSLKEGDGIYTISGGGAQGAEDKVIQHNTGGEVIIDGFEVYDFGKLYRSCGTCGDIQRKVSVSNVVAVSGSQLVGINENFGDTATIDSSVCATDVNDICATYNGTDGDEEPEEVSTGPSDYCIYTEPIAECA
9-4 : None
MVQKSLLFSLLASTALGALTKRYSFPLPESQGSETFSEPYEVAAGETFDGGMKTYGRGVECTGQVEGGEDDTVFIVQEGGTLKNAIIGTDQIEGVYCLGSCTIENVWWEKVCEDALSLKEGDGIYTISGGGAQGAEDKVIQHNTGGEVIIDGFEVYDFGKLYRSCGTCGDIQRKVSVSNVVAVSGSQLVGINENFGDTATIDSSVCATDVNDICATYNGTDGDEEPEEVSTGPSDYCIYTEPIAECA
9-2 : 插入突变

 如何对构建基因克隆的测序结果批量进行序列比对并输出结果 (qq.com)

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