#!/usr/bin/env python3
# -*- coding: utf-8 -*-
import unittest
from singlecellmultiomics.fastqProcessing.fastqIterator import FastqRecord
from singlecellmultiomics.barcodeFileParser.barcodeFileParser import BarcodeParser
from singlecellmultiomics.modularDemultiplexer.demultiplexModules.CELSeq2 import CELSeq2_c8_u6_NH
from singlecellmultiomics.modularDemultiplexer.demultiplexModules.scCHIC import SCCHIC_384w_c8_u3_cs2
from singlecellmultiomics.modularDemultiplexer.demultiplexModules.DamID import DamID2andT_SCA,DamID2_SCA
from singlecellmultiomics.modularDemultiplexer.baseDemultiplexMethods import UmiBarcodeDemuxMethod
import pkg_resources
from singlecellmultiomics.utils import reverse_complement
class TestUmiBarcodeDemux(unittest.TestCase):
def test_UmiBarcodeDemuxMethod_matching_barcode(self):
barcode_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/barcodes/')
index_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/indices/')
barcode_parser = BarcodeParser(barcode_folder, lazyLoad='*')
index_parser = BarcodeParser(index_folder, lazyLoad='*')
r1 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 1:N:0:GTGAAA',
'ATCACACACTATAGTCATTCAGGAGCAGGTTCTTCAGGTTCCCTGTAGTTGTGTGGTTTTGAGTGAGTTTTTTAAT',
'+',
'AAAAA#EEEEEEEEEEEAEEEEEEEAEEEEEEEEEEEEEEEEEE/EEEEEEEEEEEE/EEEEEEEEEEEEEEEEEE'
)
r2 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 2:N:0:GTGAAA',
'ACCCCAGATCAACGTTGGACNTCNNCNTTNTNCTCNGCACCNNNNCNNNCTTATNCNNNANNNNNNNNNNTNNGN',
'+',
'6AAAAEEAEE/AEEEEEEEE#EE##<#6E#A#EEE#EAEEA####A###EE6EE#E###E##########E##A#'
)
demux = UmiBarcodeDemuxMethod(umiRead=0,
umiStart=0,
umiLength=3,
barcodeRead=0,
barcodeStart=3,
barcodeLength=8,
barcodeFileParser=barcode_parser,
barcodeFileAlias='maya_384NLA',
indexFileParser=index_parser,
indexFileAlias='illumina_merged_ThruPlex48S_RP',
random_primer_read=None,
random_primer_length=6)
demultiplexed_record = demux.demultiplex([r1,r2])
# The barcode sequence is ACACACTA (first barcode)
self.assertEqual( demultiplexed_record[0].tags['BC'], 'ACACACTA')
self.assertEqual( demultiplexed_record[0].tags['bi'], 1) # 1 from version 0.1.12
def test_CS2_NH_matching_barcode(self):
barcode_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/barcodes/')
index_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/indices/')
barcode_parser = BarcodeParser(barcode_folder, lazyLoad='*')
index_parser = BarcodeParser(index_folder, lazyLoad='*')
seq = 'TATGAGCAATCACACACTATAGTCATTCAGGAGCAGGTTCTTCAGGTTCCCTGTAGTTGTGT'
r1 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 1:N:0:GTGAAA',
f'ATAATATCTGGGCA{seq}',
'+',
'AAAAA#EEEEEEEEEEEAEEEEEEEAEEEEEEEEEEEEEEEEEE/EEEEEEEEEEEE/EEEEEEEEEEEEEEEEEE'
)
r2 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 2:N:0:GTGAAA',
'ACCCCAGATCAACGTTGGACNTCNNCNTTNTNCTCNGCACCNNNNCNNNCTTATNCNNNANNNNNNNNNNTNNGN',
'+',
'6AAAAEEAEE/AEEEEEEEE#EE##<#6E#A#EEE#EAEEA####A###EE6EE#E###E##########E##A#'
)
demux = CELSeq2_c8_u6_NH(
barcodeFileParser=barcode_parser,
indexFileParser=index_parser)
demultiplexed_record = demux.demultiplex([r1,r2])
# The barcode sequence is ACACACTA (first barcode)
self.assertEqual( demultiplexed_record[0].tags['BC'], 'TCTGGGCA')
self.assertEqual( demultiplexed_record[0].tags['bi'], 55)
self.assertEqual( demultiplexed_record[0].tags['RX'], 'ATAATA')
self.assertEqual( demultiplexed_record[0].sequence, seq)
def construct_tchic_read(self,crx,ccb,trx,tcb,mr,linker):
seq = f'{crx}{ccb}{linker}{trx}{tcb}TTTTTTTTTTTTTTTTTTTTT{mr}'
r1 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 1:N:0:GTGAAA',
f'{seq}',
'+',
'A'*len(seq)
)
r2 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 2:N:0:GTGAAA',
reverse_complement(seq),
'+',
'A'*len(seq)
)
return r1, r2
def test_TCHIC(self):
barcode_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/barcodes/')
index_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/indices/')
barcode_parser = BarcodeParser(barcode_folder, lazyLoad='*',)
index_parser = BarcodeParser(index_folder, lazyLoad='*')
crx = 'TAT'
ccb = 'TAAGTGCT'
trx = 'CTGTTG'
tcb = 'ACAGAAGC'
mr = 'TGAGAGAGAGAGAGAGAGAGAGAGC'
linker = 'TATC'
r1,r2 = self.construct_tchic_read(crx,ccb,trx,tcb,mr,linker)
demux = SCCHIC_384w_c8_u3_cs2(
barcodeFileParser=barcode_parser,
indexFileParser=index_parser)
demultiplexed_record = demux.demultiplex([r1,r2])
# The barcode sequence is ACACACTA (first barcode)
self.assertEqual( demultiplexed_record[0].tags['BC'], ccb)
self.assertEqual( demultiplexed_record[0].tags['bi'], 225)
self.assertEqual( demultiplexed_record[0].tags['dt'], 'VASA')
self.assertEqual( demultiplexed_record[0].tags['RX'], crx)
self.assertEqual( demultiplexed_record[0].tags['rx'], trx)
self.assertEqual( demultiplexed_record[1].sequence, reverse_complement(mr)[:len(mr)-4])
r1 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 1:N:0:GTGAAA',
'GGCGACGTCCTTCACTATAGGGAGTTCTACAGTTCGACGATCCTTAAATGGTGAGTTTTTTTTTTTTTTTTTTTTTTTGACCGACGGTCCCCCCGGGACCC',
'+',
'A'*len('GGCGACGTCCTTCACTATAGGGAGTTCTACAGTTCGACGATCCTTAAATGGTGAGTTTTTTTTTTTTTTTTTTTTTTTGACCGACGGTCCCCCCGGGACCC')
)
r2 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 2:N:0:GTGAAA',
'CGATCCTTAAATGGTGAGTTTTTTTTTTTTTTTTTTTTTTTGACCGACGGTCCCCCCGGGACCCGACGGCGCGACGACGCCCGGGGCGCACTGGGGACAGT',
'+',
'A'*len('CGATCCTTAAATGGTGAGTTTTTTTTTTTTTTTTTTTTTTTGACCGACGGTCCCCCCGGGACCCGACGGCGCGACGACGCCCGGGGCGCACTGGGGACAGT')
)
demultiplexed_record = demux.demultiplex([r1,r2])
self.assertEqual( demultiplexed_record[0].tags['BC'], 'GACGTCCT')
self.assertEqual( demultiplexed_record[0].tags['bi'], 214)
self.assertEqual( demultiplexed_record[0].tags['dt'], 'VASA')
self.assertEqual( demultiplexed_record[0].tags['RX'], 'GGC')
self.assertEqual( demultiplexed_record[0].tags['rx'], 'CTTAAA')
def construct_read_pair(self, prefix, content):
seq = f'{prefix}{content}'
r1 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 1:N:0:GTGAAA',
f'{seq}',
'+',
'A'*len(seq)
)
r2 = FastqRecord(
'@NS500414:628:H7YVNBGXC:1:11101:15963:1046 2:N:0:GTGAAA',
reverse_complement(seq),
'+',
'A'*len(seq)
)
return r1, r2
def test_DAMID(self):
barcode_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/barcodes/')
index_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/indices/')
barcode_parser = BarcodeParser(barcode_folder, lazyLoad='*',)
index_parser = BarcodeParser(index_folder, lazyLoad='*')
# First single cell format without overhang:
# EG: DamID2_BC_001 3-TGCA-3-TATG
first_barcode, second_barcode = 'TGCA', 'TATG'
first_umi = 'ACT'
second_umi='CTC'
read_contents = 'rrrrrr'
r1,r2 = self.construct_read_pair(f'{first_umi}{first_barcode}{second_umi}{second_barcode}',read_contents)
damid_demux = DamID2_SCA(barcodeFileParser=barcode_parser,
second_barcode_len=4,
indexFileParser=index_parser,
barcode_alias='DamID2_scattered_8bp',
)
demultiplexed_record = damid_demux.demultiplex([r1,r2])
self.assertEqual( demultiplexed_record[0].tags['BC'], first_barcode+second_barcode)
self.assertEqual( demultiplexed_record[0].tags['bi'], 1)
self.assertEqual( demultiplexed_record[0].tags['RX'], first_umi+second_umi)
self.assertEqual( demultiplexed_record[0].sequence, read_contents)
combined_demux = DamID2andT_SCA(
barcodeFileParser=barcode_parser,
indexFileParser=index_parser)
# The internal DamID demux of the combined protocol should return the same result
demultiplexed_record = combined_demux.damid_demux.demultiplex([r1,r2])
self.assertEqual( demultiplexed_record[0].tags['BC'], first_barcode+second_barcode)
self.assertEqual( demultiplexed_record[0].tags['bi'], 1)
self.assertEqual( demultiplexed_record[0].tags['RX'], first_umi+second_umi)
self.assertEqual( demultiplexed_record[0].sequence, read_contents)
demultiplexed_record = combined_demux.demultiplex([r1,r2])
self.assertEqual( demultiplexed_record[0].tags['BC'], first_barcode+second_barcode)
self.assertEqual( demultiplexed_record[0].tags['bi'], 1)
self.assertEqual( demultiplexed_record[0].tags['dt'], 'DamID')
self.assertEqual( demultiplexed_record[0].tags['RX'], first_umi+second_umi)
self.assertEqual( demultiplexed_record[0].sequence, read_contents)
def test_3DEC_UmiBarcodeDemuxMethod_matching_barcode(self):
barcode_folder = pkg_resources.resource_filename('singlecellmultiomics','modularDemultiplexer/barcodes/')
barcode_parser = BarcodeParser(barcode_folder,lazyLoad='*')
r1 = FastqRecord(
'@Cluster_s_1_1101_1000',
'ATCACACACTATAGTCATTCAGGAGCAGGTTCTTCAGGTTCCCTGTAGTTGTGTGGTTTTGAGTGAGTTTTTTAAT',
'+',
'AAAAA#EEEEEEEEEEEAEEEEEEEAEEEEEEEEEEEEEEEEEE/EEEEEEEEEEEE/EEEEEEEEEEEEEEEEEE'
)
r2 = FastqRecord(
'@Cluster_s_1_1101_1002',
'ACCCCAGATCAACGTTGGACNTCNNCNTTNTNCTCNGCACCNNNNCNNNCTTATNCNNNANNNNNNNNNNTNNGN',
'+',
'6AAAAEEAEE/AEEEEEEEE#EE##<#6E#A#EEE#EAEEA####A###EE6EE#E###E##########E##A#'
)
demux = UmiBarcodeDemuxMethod(umiRead=0,
umiStart=0,
umiLength=3,
barcodeRead=0,
barcodeStart=3,
barcodeLength=8,
barcodeFileParser=barcode_parser,
barcodeFileAlias='maya_384NLA',
indexFileParser=None,
indexFileAlias='illumina_merged_ThruPlex48S_RP',
random_primer_read=None,
random_primer_length=6)
demultiplexed_record = demux.demultiplex([r1,r2])
# The barcode sequence is ACACACTA (first barcode)
self.assertEqual( demultiplexed_record[0].tags['BC'], 'ACACACTA')
self.assertEqual( demultiplexed_record[0].tags['bi'], 1)
if __name__ == '__main__':
unittest.main()
Datasets
Models
