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SRSF6 / Serine/arginine-rich splicing factor 6 · Western blot design guide
Design a Western Blot for SRSF6
Source-linked SRSF6 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-SRSF6 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Evidence assembled September 2026 · For research use; verify linked source records and product datasheet before use
SRSF6 Western Blot Experimental Design Guide
Expected bands, source-linked protocol options, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~39.6 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Adipose tissue (IHC candidate; verify WB) +4 more | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 3 isoform(s) |
Section 1
Source-Linked SRSF6 Western Blot Protocol Options
The M04489 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | K562 cell lysate (catalog M04489) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | M04489; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Species-matched HRP conjugate at validated dilution (standard starting point) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL; bracket exposures to avoid saturation (standard starting point) |
Section 2
What Is the Expected SRSF6 Western Blot Band Size?
SRSF6 is predicted at 39.6 kDa; isoforms and phosphorylation could affect migration, but no empirical band size or visible shift is established.
| Band near 39.6 kDa | consistent with the predicted mass of SRSF6; confirm identity with controls |
| Several bands at different positions | could reflect SRP55-1, SRP55-2, and SRP55-3; their separation is unverified |
| Doublet near the predicted position | could reflect different phosphorylation states; a mobility effect is unverified |
| Stronger band in nuclear than cytosolic fraction | consistent with SRSF6 localization to the nucleus and nuclear speckles |
💡Expected SRSF6 appearanceSRSF6 has a predicted mass of 39.6 kDa, but no empirical band size is supplied; use knockdown or another band-identity control to assess any detected band.
| UniProt predicted mass | sets a 39.6 kDa sequence-based reference, not a measured migration position |
| SRP55-1 isoform | may differ in apparent size from other isoforms; its relative position is unknown |
| SRP55-2 isoform | may differ in apparent size from other isoforms; its relative position is unknown |
| SRP55-3 isoform | may differ in apparent size from other isoforms; its relative position is unknown |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | poor recovery of nuclear SRSF6 is possible | check a nuclear fraction, loading, and a positive control |
| Band higher than expected | phosphorylation may affect mobility, but no shift is established | compare phosphatase-treated and untreated samples and verify band identity |
| Band lower than expected | an alternative isoform is possible; isoform masses are unavailable | confirm identity by SRSF6 knockdown and compare isoform expression |
| Multiple bands | SRP55 isoforms or distinct phosphorylation states are possible | compare SRSF6 knockdown and phosphatase-treated samples |
| Weak or no signal | nuclear SRSF6 may be poorly represented in the tested fraction | check nuclear enrichment and a positive control |
Experiment evidence
Western blot images showing SRSF6 band
M04489 · SRSF6
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for SRSF6 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for SRSF6 in Western blot, you can use adipose tissue lysate, which has high HPA expression.
Positive control: Adipose tissue (IHC candidate; verify WB)
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: No tissue is reported as not detected, so use siRNA knockdown or a KO line as the negative control.
HPA tissue expression evidence for SRSF6
Comprehensive Human Protein Atlas IHC scoring per tissue. Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Higher expression tissues · candidate positive controls from IHC
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | adipocytes | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | High | Protein (IHC) | HPA → |
| Appendix | glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | adipocytes | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Liver | cholangiocytes | Medium | Protein (IHC) | HPA → |
| Lymph node | non-germinal center cells | Medium | Protein (IHC) | HPA → |
| Ovary | ovarian stroma cells | Medium | Protein (IHC) | HPA → |
| Parathyroid gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Skeletal muscle | myocytes | Medium | Protein (IHC) | HPA → |
Section 3
Advanced SRSF6 Western Blot Tips
Deeper troubleshooting and optimisation questions for SRSF6, answered from its protein features.
How should SRSF6 band migration be interpreted?
Band shift · Use the separately labelled calculated mass and catalog-observed evidence above. A sequence annotation does not establish an observed migration shift. Verify target identity with orthogonal controls.
How might SRSF6 isoforms affect antibody detection?
Isoforms · UniProt lists SRP55-1, SRP55-2, and SRP55-3. In SRP55-2, residues 86–135 are replaced and 136–344 are missing; in SRP55-3, residues 313–344 are replaced. Check whether the antibody epitope is retained in the isoform you intend to detect.
Which SRSF6 phosphorylation sites are relevant to band interpretation?
PTM · UniProt lists phosphoserines at positions 45, 81, 84, 297, 299, 303, 314, and 316. These are UniProt canonical sequence coordinates; antibody or paper numbering may differ. Their presence alone does not establish a visible band shift.
Yes. UniProt lists N6-acetyllysine at position 165 in canonical sequence coordinates. SRP55-2 lacks residues 136–344, so that annotated site is absent from that isoform. Acetylation alone does not establish a detectable band shift.
Yes. UniProt lists N6-acetyllysine at position 165 in canonical sequence coordinates. SRP55-2 lacks residues 136–344, so that annotated site is absent from that isoform. Acetylation alone does not establish a detectable band shift.
Does this guide establish induction of SRSF6?
Induction · No general induction response is established by this guide. A pathway or gene-set association is not evidence of induction in a particular specimen. Verify the relevant treatment and control in a target-specific experiment.
How should transfer be checked for SRSF6?
Transfer · Standard workflow guidance: verify transfer efficiency for the intended target size before interpreting a weak signal. Use total-protein assessment and optimize transfer for the membrane, gel and apparatus; the labelled catalog values take precedence.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the M04489 datasheet where specified. Otherwise compare 5% milk or 5% BSA in TBST; for a phospho-specific assay start with BSA. Optimize background and specific signal with matched controls.
How should SRSF6 be quantified across samples?
Quantitation · SRSF6 is annotated in the nucleus and nuclear speckles. Compare the same sample fraction and preparation across conditions, and normalize within that fraction. If using nuclear extracts, keep extraction consistent so differences in recovery are not mistaken for differences in SRSF6 abundance.
Should SRSF6 run at its predicted 39.6 kDa?
Interpretation · 39.6 kDa is the predicted mass, but no observed band position is supplied. The listed modifications and isoforms do not establish a visible shift or explain a difference from 39.6 kDa. Use the prediction as a reference, not a required band position.
UniProt identifies phosphoserine 303 as modified by DYRK1A and reports an interaction between SRSF6 and DYRK1A. If studying DYRK1A conditions, distinguish changes in phosphorylation from changes in total SRSF6; the supplied features do not establish an induced change in abundance.
Check antibody epitope coverage against the three isoforms, especially the altered regions at 86–135, 136–344, and 313–344. Consider the listed phosphorylation and acetylation sites as possibilities, but do not assign an extra band to a specific isoform or modification from position alone.
UniProt identifies phosphoserine 303 as modified by DYRK1A and reports an interaction between SRSF6 and DYRK1A. If studying DYRK1A conditions, distinguish changes in phosphorylation from changes in total SRSF6; the supplied features do not establish an induced change in abundance.
Check antibody epitope coverage against the three isoforms, especially the altered regions at 86–135, 136–344, and 313–344. Consider the listed phosphorylation and acetylation sites as possibilities, but do not assign an extra band to a specific isoform or modification from position alone.
Boster reagents
SRSF6 Western Blot Antibodies
Catalog antibodies with Western blot application and product-specific WB images. Evaluate suitability with the reported sample, controls and experimental conditions.
Real WB data
Anti-SR protein repeat Rabbit Monoclonal Antibody
Cat # M04489
The listed anti-SRSF6 antibody, M04489, is a rabbit monoclonal with reported human reactivity. Its Western blot image shows SR protein repeat expression in K562 cell lysate; the supplied evidence covers this tested context only.
Which to pick: M04489 is the only listed option. Choose it for a human K562 Western blot context supported by the reported image; confirm suitability separately for other samples or conditions.
Source: BosterBio SRSF6 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q13247.
- Human Protein Atlas. SRSF6 tissue expression.
- PMC7551351 — target-verified WB comparison
- PMC7441155 — target-verified WB comparison
- PMC9881134 — target-verified WB comparison