All primers used in high throughput are labelled with 6-nucleotide barcode sequences in the 5’ end of the primer sequences listed in the table below.

  For the variable region of IgG and IgA heavy chains, two degenerate primers for mice (mH1 and mH2) are used together as one forward primer, the reverse primers on constant region ensure the specificity of IgG or IgA heavy chain class.

Primer name

Sequence

mVK-Forward

GACATTCAGCTGACCCAGTCTCCA

mH1-Forward

SARGTNMAGCTGSAGSAGTC

mH2-Forward

SARGTNMAGCTGSAGSAGTCWGG

mVK-Reverse

AGNTTGGTTCCACCRCCGAACG

mVG-Reverse

GATAGACHGATGGGGSTGTYGTT

mVA-reverse

TGGTGGGATTTCTCGCAGAC


No. FW Primer FW Barcode Nucleotides RW Primer RW Barcode Nucleotides Sample Name
1 mVK-BC01F barcode1 ATCACG mVK-BC10R barcode10 TAGCTT μMT-Spleen-PBS-2-Igk
2 mVK-BC01F barcode1 ATCACG mVK-BC11R barcode11 GGCTAC μMT-Liver-PBS-1-Igk
3 mVK-BC01F barcode1 ATCACG mVK-BC12R barcode12 CTTGTA μMT-Liver-PBS-2-Igk
4 mVK-BC01F barcode1 ATCACG mVK-BC14R barcode14 AGTTCC μMT-Liver-PBS-4-Igk
5 mVK-BC01F barcode1 ATCACG mVK-BC15R barcode15 ATGTCA μMT-Liver-NP-6-Igk
6 mVK-BC01F barcode1 ATCACG mVK-BC16R barcode16 CCGTCC μMT-Liver-NP-7-Igk
7 mVK-BC01F barcode1 ATCACG mVK-BC18R barcode18 GTCCGC μMT-Liver-NP-9-Igk
8 mVK-BC01F barcode1 ATCACG mVK-BC20R barcode20 GTGGCC μMT-Lung-PBS-1-Igk
9 mVK-BC01F barcode1 ATCACG mVK-BC21R barcode21 GTTTCG μMT-Lung-PBS-2-Igk
10 mVK-BC01F barcode1 ATCACG mVK-BC22R barcode22 CGTACG μMT-Lung-PBS-3-Igk
11 mVK-BC01F barcode1 ATCACG mVK-BC23R barcode23 GAGTGG μMT-Lung-PBS-4-Igk
12 mVK-BC01F barcode1 ATCACG mVK-BC24R barcode24 GGTAGC μMT-Lung-NP-6-Igk
13 mVK-BC01F barcode1 ATCACG mVK-BC27R barcode27 ATTCCT μMT-Lung-NP-10-Igk
14 mVK-BC01F barcode1 ATCACG mVK-BC28R barcode28 CAAAAG μMT-Kidney-PBS-1-Igk
15 mVK-BC01F barcode1 ATCACG mVK-BC29R barcode29 CAACTA μMT-Kidney-PBS-2-Igk
16 mVK-BC01F barcode1 ATCACG mVK-BC32R barcode32 CACTCA μMT-Kidney-NP-6-Igk
17 mVK-BC01F barcode1 ATCACG mVK-BC34R barcode34 CATGGC μMT-Kidney-NP-9-Igk
18 mVK-BC01F barcode1 ATCACG mVK-BC37R barcode37 CGGAAT BALBc-Kidney-PBS-5-Igk
19 mVK-BC01F barcode1 ATCACG mVK-BC38R barcode38 CTAGCT BALBc-Kidney-NP-6-Igk
20 mVK-BC01F barcode1 ATCACG mVK-BC39R barcode39 CTATAC BALBc-Kidney-NP-7-Igk
21 mVK-BC01F barcode1 ATCACG mVK-BC40R barcode40 CTCAGA BALBc-Kidney-NP-9-Igk
22 mVK-BC01F barcode1 ATCACG mVK-BC41R barcode41 GACGAC BALBc-Lung-PBS-4-Igk
23 mVK-BC01F barcode1 ATCACG mVK-BC42R barcode42 TAATCG BALBc-Lung-PBS-5-Igk
24 mVK-BC01F barcode1 ATCACG mVK-BC43R barcode43 TACAGC BALBc-Lung-NP-6-Igk
25 mVK-BC01F barcode1 ATCACG mVK-BC44R barcode44 TATAAT BALBc-Lung-NP-8-Igk
26 mVK-BC01F barcode1 ATCACG mVK-BC45R barcode45 TCATTC μMT-Lung-NP-6-Igk
27 mVK-BC01F barcode1 ATCACG mVK-BC46R barcode46 TCCCGA μMT-Lung-NP-12-Igk
28 mVK-BC01F barcode1 ATCACG mVK-BC47R barcode47 TCGAAG BALBc-Spleen-PBS-Igk
29 mVK-BC02F barcode2 CGATGT mVK-BC10R barcode10 TAGCTT μMT_CD3-T_Sort1-Igk
30 mVK-BC02F barcode2 CGATGT mVK-BC11R barcode11 GGCTAC μMT_CD3-T_Sort2-Igk
31 mVK-BC02F barcode2 CGATGT mVK-BC12R barcode12 CTTGTA μMT_CD3-T_Sort3-Igk
32 mVK-BC02F barcode2 CGATGT mVK-BC13R Barcode13 AGTCAA μMT_CD4-T_PBS4-Igk
33 mVK-BC02F barcode2 CGATGT mVK-BC14R barcode14 AGTTCC μMT_CD8-T_PBS4-Igk
34 mVK-BC02F barcode2 CGATGT mVK-BC15R barcode15 ATGTCA μMT_CD4-T_NP7-Igk
35 mVK-BC02F barcode2 CGATGT mVK-BC16R barcode16 CCGTCC μMT_CD8-T_NP7-Igk
36 mVK-BC02F barcode2 CGATGT mVK-BC17R barcode17 GTAGAG μMT_CD4-T_NP8-Igk
37 mVK-BC02F barcode2 CGATGT mVK-BC18R barcode18 GTCCGC μMT_CD8-T_NP8-Igk
38 mVK-BC02F barcode2 CGATGT mVK-BC19R barcode19 GTGAAA μMT_CD4-T_NP12-Igk
39 mVK-BC02F barcode2 CGATGT mVK-BC20R barcode20 GTGGCC μMT_CD8-T_NP12-Igk
40 mVK-BC02F barcode2 CGATGT mVK-BC21R barcode21 GTTTCG BALBc_CD4-T_PBS-Igk
41 mVK-BC02F barcode2 CGATGT mVK-BC22R barcode22 CGTACG BALBc_CD8-T_PBS-Igk
42 mVK-BC02F barcode2 CGATGT mVK-BC23R barcode23 GAGTGG BALBc_B_PBS-Igk
43 mH1/2-BC04F barcode4 TGACCA mVG-BC11R barcode11 GGCTAC μMT_Liver_PBS2_Igγ
44 mH1/2-BC04F barcode4 TGACCA mVG-BC12R barcode12 CTTGTA μMT_Liver_PBS3_Igγ
45 mH1/2-BC04F barcode4 TGACCA mVG-BC13R Barcode13 AGTCAA μMT_Liver_PBS4_Igγ
46 mH1/2-BC04F barcode4 TGACCA mVG-BC14R barcode14 AGTTCC μMT_Liver_NP6_Igγ
47 mH1/2-BC04F barcode4 TGACCA mVG-BC15R barcode15 ATGTCA μMT_Liver_NP7_Igγ
48 mH1/2-BC04F barcode4 TGACCA mVG-BC16R barcode16 CCGTCC μMT_Liver_NP8_Igγ
49 mH1/2-BC04F barcode4 TGACCA mVG-BC17R barcode17 GTAGAG μMT_Liver_NP9_Igγ
50 mH1/2-BC04F barcode4 TGACCA mVG-BC18R barcode18 GTCCGC μMT_Lung_PBS1_Igγ
51 mH1/2-BC04F barcode4 TGACCA mVG-BC21R barcode21 GTTTCG μMT_Lung_PBS4_Igγ
52 mH1/2-BC04F barcode4 TGACCA mVG-BC23R barcode23 GAGTGG μMT_Lung_NP7_Igγ
53 mH1/2-BC04F barcode4 TGACCA mVG-BC26R barcode26 ATGAGC μMT_Kidney_PBS1_Igγ
54 mH1/2-BC05F barcode5 ACAGTG mVG-BC10R barcode10 TAGCTT μMT_Kidney_NP6_Igγ
55 mH1/2-BC05F barcode5 ACAGTG mVG-BC11R barcode11 GGCTAC μMT_Kidney_NP7_Igγ
56 mH1/2-BC05F barcode5 ACAGTG mVG-BC17R barcode17 GTAGAG μMT_Spleen_PBS2_Igγ
57 mH1/2-BC05F barcode5 ACAGTG mVG-BC18R barcode18 GTCCGC BALBc_Spleen_PBS_Igγ
58 mH1/2-BC05F barcode5 ACAGTG mVG-BC21R barcode21 GTTTCG BALBc_Kidney_PBS4-Sorted_Igγ
59 mH1/2-BC05F barcode5 ACAGTG mVG-BC23R barcode23 GAGTGG BALBc_Kidney_NP7-Sorted_Igγ
60 mH1/2-BC05F barcode5 ACAGTG mVG-BC24R barcode24 GGTAGC BALBc_Kidney_NP8-Sorted_Igγ
61 mH1/2-BC05F barcode5 ACAGTG mVG-BC27R barcode27 ATTCCT BALBc_Lung_PBS1-Sorted_Igγ
62 mH1/2-BC05F barcode5 ACAGTG mVG-BC28R barcode28 CAAAAG BALBc_Lung_PBS3-Sorted_Igγ
63 mH1/2-BC05F barcode5 ACAGTG mVG-BC29R barcode29 CAACTA BALBc_Lung_NP6-Sorted_Igγ
64 mH1/2-BC06F barcode6 GCCAAT mVG-BC11R barcode11 GGCTAC Lung_NP10-Sorted_Igγ
65 mH1/2-BC06F barcode6 GCCAAT mVG-BC12R barcode12 CTTGTA BALBc_CD4+T_PBS-Sorted_Igγ
66 mH1/2-BC06F barcode6 GCCAAT mVG-BC13R Barcode13 AGTCAA BALBc_CD8+T_PBS-Sorted_Igγ
67 mH1/2-BC06F barcode6 GCCAAT mVG-BC14R barcode14 AGTTCC BALBc_B_PBS-Sorted_Igγ
68 mH1/2-BC06F barcode6 GCCAAT mVG-BC15R barcode15 ATGTCA BALBc_CD4+T_NP7-Sorted_Igγ
69 mH1/2-BC06F barcode6 GCCAAT mVG-BC17R barcode17 GTAGAG BALBc_Macrophage_PBS1-Sorted_Igγ
70 mH1/2-BC06F barcode6 GCCAAT mVG-BC18R barcode18 GTCCGC BALBc_Macrophage_PBS2-Sorted_Igγ
71 mH1/2-BC06F barcode6 GCCAAT mVG-BC19R barcode19 GTGAAA BALBc_Macrophage_PBS3-Sorted_Igγ
72 mH1/2-BC06F barcode6 GCCAAT mVG-BC20R barcode20 GTGGCC μMT_CD4+T_PBS4_Igγ
73 mH1/2-BC06F barcode6 GCCAAT mVG-BC21R barcode21 GTTTCG μMT_CD8+T_PBS4_Igγ
74 mH1/2-BC06F barcode6 GCCAAT mVG-BC22R barcode22 CGTACG μMT_CD4+T_NP8_Igγ
75 mH1/2-BC06F barcode6 GCCAAT mVG-BC23R barcode23 GAGTGG μMT_CD8+T_NP8_Igγ
76 mH1/2-BC06F barcode6 GCCAAT mVG-BC24R barcode24 GGTAGC μMT_CD4+T_NP12_Igγ
77 mH1/2-BC06F barcode6 GCCAAT mVG-BC25R barcode25 ACTGAT μMT_CD8+T_NP12_Igγ
78 mH1/2-BC04F barcode4 TGACCA mVA-BC30R barcode30 CACCGG μMT_Liver_PBS1_Igα
79 mH1/2-BC04F barcode4 TGACCA mVA-BC31R barcode31 CACGAT μMT_Liver_PBS2_Igα
80 mH1/2-BC04F barcode4 TGACCA mVA-BC32R barcode32 CACTCA μMT_Liver_PBS3_Igα
81 mH1/2-BC04F barcode4 TGACCA mVA-BC33R barcode33 CAGGCG μMT_Liver_PBS4_Igα
82 mH1/2-BC04F barcode4 TGACCA mVA-BC34R barcode34 CATGGC μMT_Liver_NP6_Igα
83 mH1/2-BC04F barcode4 TGACCA mVA-BC35R barcode35 CATTTT μMT_Liver_NP7_Igα
84 mH1/2-BC04F barcode4 TGACCA mVA-BC37R barcode37 CGGAAT μMT_Liver_NP9_Igα
85 mH1/2-BC04F barcode4 TGACCA mVA-BC38R barcode38 CTAGCT μMT_Lung_PBS1_Igα
86 mH1/2-BC04F barcode4 TGACCA mVA-BC39R barcode39 CTATAC μMT_Lung_PBS2_Igα
87 mH1/2-BC04F barcode4 TGACCA mVA-BC40R barcode40 CTCAGA μMT_Lung_PBS3_Igα
88 mH1/2-BC04F barcode4 TGACCA mVA-BC41R barcode41 GACGAC μMT_Lung_PBS4_Igα
89 mH1/2-BC04F barcode4 TGACCA mVA-BC42R barcode42 TAATCG μMT_Lung_NP6_Igα
90 mH1/2-BC04F barcode4 TGACCA mVA-BC46R barcode46 TCCCGA μMT_Kidney_PBS1_Igα
91 mH1/2-BC04F barcode4 TGACCA mVA-BC47R barcode47 TCGAAG μMT_Kidney_PBS2_Igα
92 mH1/2-BC05F barcode5 ACAGTG mVA-BC33R barcode33 CAGGCG μMT_Kidney_NP8_Igα
93 mH1/2-BC05F barcode5 ACAGTG mVA-BC34R barcode34 CATGGC μMT_Kidney_NP9_Igα
94 mH1/2-BC05F barcode5 ACAGTG mVA-BC35R barcode35 CATTTT μMT_Liver_NP10_Igα
95 mH1/2-BC05F barcode5 ACAGTG mVA-BC37R barcode37 CGGAAT μMT_Kidney_NP10_Igα
96 mH1/2-BC05F barcode5 ACAGTG mVA-BC39R barcode39 CTATAC BALBc_Spleen_PBS_Igα
97 mH1/2-BC05F barcode5 ACAGTG mVA-BC40R barcode40 CTCAGA BALBc_Kidney_PBS2-Sorted_Igα
98 mH1/2-BC05F barcode5 ACAGTG mVA-BC41R barcode41 GACGAC BALBc_Kidney_PBS3-Sorted_Igα
99 mH1/2-BC05F barcode5 ACAGTG mVA-BC42R barcode42 TAATCG BALBc_Kidney_PBS4-Sorted_Igα
100 mH1/2-BC05F barcode5 ACAGTG mVA-BC43R barcode43 TACAGC BALBc_Kidney_PBS5-Sorted_Igα
101 mH1/2-BC05F barcode5 ACAGTG mVA-BC44R barcode44 TATAAT BALBc_Kidney_NP7-Sorted_Igα
102 mH1/2-BC05F barcode5 ACAGTG mVA-BC45R barcode45 TCATTC BALBc_Kidney_NP7-Sorted_Igα
103 mH1/2-BC05F barcode5 ACAGTG mVA-BC46R barcode46 TCCCGA BALBc_Kidney_NP9-Sorted_Igα
104 mH1/2-BC05F barcode5 ACAGTG mVA-BC48R barcode48 TCGGCA μMT_Lung_PBS1-Sorted_Igα
105 mH1/2-BC06F barcode6 GCCAAT mVA-BC30R barcode30 CACCGG μMT_Lung_PBS3-Sorted_Igα
106 mH1/2-BC06F barcode6 GCCAAT mVA-BC31R barcode31 CACGAT μMT_Lung_NP6-Sorted_Igα
107 mH1/2-BC06F barcode6 GCCAAT mVA-BC33R barcode33 CAGGCG μMT_Lung_NP10-Sorted_Igα
108 mH1/2-BC06F barcode6 GCCAAT mVA-BC34R barcode34 CATGGC BALBc_CD4+T_PBS_Igα
109 mH1/2-BC06F barcode6 GCCAAT mVA-BC35R barcode35 CATTTT BALBc_CD8+T_PBS_Igα
110 mH1/2-BC06F barcode6 GCCAAT mVA-BC36R barcode36 CCAACA BALBc_B_PBS_Igα
111 mH1/2-BC06F barcode6 GCCAAT mVA-BC37R barcode37 CGGAAT BALBc_CD4+T_NP7_Igα
112 mH1/2-BC06F barcode6 GCCAAT mVA-BC38R barcode38 CTAGCT BALBc_CD8+T_NP7_Igα
113 mH1/2-BC06F barcode6 GCCAAT mVA-BC39R barcode39 CTATAC BALBc_Macrophage_PBS1_Igα
114 mH1/2-BC06F barcode6 GCCAAT mVA-BC40R barcode40 CTCAGA BALBc_Macrophage_PBS2_Igα
115 mH1/2-BC06F barcode6 GCCAAT mVA-BC41R barcode41 GACGAC BALBc_Macrophage_PBS3_Igα
116 mH1/2-BC06F barcode6 GCCAAT mVA-BC43R barcode43 TACAGC μMT_CD8+T_PBS4_Igα
117 mH1/2-BC06F barcode6 GCCAAT mVA-BC44R barcode44 TATAAT μMT_CD4+T_NP8_Igα
118 mH1/2-BC06F barcode6 GCCAAT mVA-BC45R barcode45 TCATTC μMT_CD8+T_NP8_Igα
119 mH1/2-BC06F barcode6 GCCAAT mVA-BC46R barcode46 TCCCGA μMT_CD4+T_NP12_Igα
120 mH1/2-BC06F barcode6 GCCAAT mVA-BC47R barcode47 TCGAAG μMT_CD8+T_NP12_Igα


Homo Sapiens (Human) Primers (used in Sanger sequencing)


Ig heavy chain
Nest PCR or half-nest PCR

First round:
VH1 (5'-GAGGTGCAGCTCGAGGAGTCTGGG-3'),
VH2 (5'-CAGGTGCAGCTCGAGCAGTCTGGG-3'),
VH3 (5'-CAGGTACAGCTCGAGCAGTCAGG-3'),
VH4 (5'-CAGGTGCAGCTGCTCGAGTCGGG-3'),
Cμ-CH1, 5'-ACGCTGC TCGTATCCGACGGG-3'
Or CγCH1, 5'-ACGCTGCTCGTATCCGACGGG-3'

Second round:
FR2 coupled with JH primer was used
FR2: 5’-TGG(A/G)TCCG(A/C/G)CAG(G/C)C(T/C)CC(A/C/G/T)GG-3’;
TGGRTCCGVCAGSCYCCNGG
JH: 5’-AACTGCAGAGGAGACGGTGACC-3’
or JH2: 5’-GTGACCAGGGTNCCTTGGCCCCAAG-3'

Ig kappa chain

Vκ1 (5’-GACATCGAGCTCACCCAGTCTCC-3’) and
Vκ 2 (5’-GAAATTGAGCTCACGCAGTCTCCA-3’)

Cκ2 primer (Cκ2: 5’-CGGGAAGATGAAGACAGATGGTGC-3’)
Half-nest PCR
Cκ1 primer (5’-TGGTGCAGCCACAGTTCGTTT-3’)
Or
JK1/2/3/4 : 5’-CTTACGTTTGATCTCCACCTTGGTCCC-3’
JK5 : 5’-CTTACGTTTAATCTCCAGTCGTGTCCC-3’